CN112778299B

CN112778299B - Piperazine ureido derivatives, preparation method thereof and application thereof in medicines

Info

Publication number: CN112778299B
Application number: CN202011180642.6A
Authority: CN
Inventors: 李心; 张志高; 陈凌翔; 谢志超; 贺峰; 陶维康
Original assignee: Jiangsu Hengrui Medicine Co Ltd; Shanghai Hengrui Pharmaceutical Co Ltd
Current assignee: Jiangsu Hengrui Medicine Co Ltd; Shanghai Hengrui Pharmaceutical Co Ltd
Priority date: 2019-11-04
Filing date: 2020-10-29
Publication date: 2023-07-14
Anticipated expiration: 2040-10-29
Also published as: CN112778299A

Abstract

The present disclosure relates to piperazinyl ureido derivatives, methods of preparing the same, and their use in medicine. In particular, the present disclosure relates to piperazinyl ureido derivatives of general formula (I), methods for their preparation, pharmaceutical compositions containing the derivatives, and their use as capsid protein inhibitors, particularly in the prevention and/or treatment of hepatitis b, influenza, herpes, aids and other diseases. Wherein each group in the general formula (I) is defined as the specification.

Description

Piperazine ureido derivatives, preparation method thereof and application thereof in medicines

Technical Field

The present disclosure relates to piperazine ureido derivatives, a preparation method thereof and application thereof in medicine, and belongs to the field of medicine. In particular, the present disclosure relates to piperazinyl ureido derivatives of general formula (I), methods for their preparation, pharmaceutical compositions containing the derivatives, and their use as capsid protein inhibitors, particularly in the prevention and/or treatment of hepatitis b, influenza, herpes, aids and other diseases.

Background

Chronic Hepatitis B Virus (HBV) infection has been a global health problem, and about 20 million people worldwide have been reported by the world health organization to be infected with HBV, of which 2.4 million people are chronically HBV infected, and about 65 tens of thousands die each year due to liver failure, cirrhosis and hepatocellular carcinoma (HCC) caused by HBV infection. The proportion caused by HBV infection in global cirrhosis and HCC patients was 30% and 45%, respectively. Although prophylactic HBV vaccines can be used, chronic HBV infection has become a worldwide medical problem due to the lack of effective therapeutics.

Currently, there are two main classes of drugs for the treatment of chronic HBV: alpha-interferon formulations (e.g., pegylated alpha-interferon) and nucleoside analogs that inhibit HBV DNA polymerase (e.g., lamivudine, adefovir, etc.). However, interferon-type formulations have serious side effects, are poorly tolerated, and only a small fraction of patients may have sustained clinical responses to interferon treatment (Lancet.2005Jan 8-14;365 (9454): 123-9.; N Engl JMed.2005Jun 30;352 (26): 2682-95.; hepatology.2009May;49 (5 Suppl): S103-11). As competitive inhibitors of reverse transcriptase, nucleoside analogue drugs exert antiviral effects by blocking the synthesis of HBV DNA strands. However, the existing nucleoside analogue drugs also have the problems of poor curative effect on drug-resistant strains and the like due to the fact that reverse transcriptase is induced to generate drug-resistant mutation. In addition, such drugs often have difficulty in thoroughly eliminating HBV infection, and cannot be radically cured even if taken for a long period of time; once stopped, severe withdrawal may also result, and thus lifelong medications are often required (hepatology. 2009May;49 (5 Suppl): S112-21). Therefore, there is an urgent need to develop new, safe and effective drugs for chronic hepatitis b.

The reason for the low cure rate of chronic HBV infection has a great relationship with the characteristics of Hepatitis B Virus (HBV), an enveloped, partially double-stranded DNA (dsDNA) virus of Hepadnaviridae family (hepadnavidae). The outermost layer of the mature HBV virus particles is envelope protein, and is wrapped by HBV Nucleocapsid (Nucleocapid). The nucleocapsid, also called Core particle, is composed of Capsid protein (Capid protein), HBV Relaxed circular DNA (rcDNA) and HBV reverse transcriptase bound to the 5' end of the negative strand of rcDNA. At the time of infection, rcDNA is converted into covalently closed circular DNA (cccDNA) in the host cell nucleus, which serves as a HBV replication template. An important step in HBV replication is capsid assembly (encapsation). The Pregenomic RNA (pgRNA) transcribed from cccDNA needs to be encapsulated in capsid protein together with HBV reverse transcriptase to complete the assembly step to enable subsequent reverse transcription. Prior to reverse transcription, HBV reverse transcriptase, pgRNA needs to be correctly encapsulated by capsid protein. Thus, blocking capsid protein assembly, or accelerating capsid protein degradation, will block capsid assembly processes, thereby affecting viral replication. In addition, the N-terminal 149 amino acid residues (Cp 149) constituting the core protein dimerization motif (Dimerization motif) and the assembly domain (Assemblelydomain) are devoid of human protein homology sequences. Therefore, capsid protein assembly inhibitors are considered as new targets for anti-hepatitis b drug development. Because of the different mechanism of action from traditional antiviral drugs, combination therapies of a capsid protein inhibitor and a DNA polymerase inhibitor should synergistically inhibit HBV replication and prevent the development of drug resistance, providing safer and more effective treatment of chronic hepatitis b infection.

Currently, there are two main classes of drugs for capsid inhibitors: heteroaryl dihydropyrimidines (HAPs) and phenylacrylamides, such as GLS-4, NVR-3778, and the like. Related patents are as follows: WO2001068642, WO2014029193, WO2015011281, WO2016016196, WO 2017076791 and WO2016113273 etc., but compounds directed to this target are mostly in the clinical stage of research and no marketed drugs are present. Thus, there is a continuing need to develop drugs for capsid inhibitors.

Disclosure of Invention

The object of the present disclosure is to provide a compound represented by the general formula (I) or a tautomer, racemate, enantiomer, diastereomer or a mixture thereof, or a pharmaceutically acceptable salt thereof,

wherein:

ring a is aryl and heteroaryl;

L ¹ selected from alkylene, -O-and-S-;

R ¹ selected from the group consisting of hydrogen, halogen, alkyl, cyano, amino, nitro, hydroxyl, haloalkyl, hydroxyalkyl, cycloalkyl, heterocyclyl, aryl, and heteroaryl;

R ² the same or different and are each independently selected from the group consisting of hydrogen, halogen, alkyl, cyano, amino, nitro, hydroxyl, haloalkyl, hydroxyalkyl, cycloalkyl, and heterocyclyl;

R ³ selected from the group consisting of a hydrogen atom, an alkyl group, a haloalkyl group, an alkoxy group, a haloalkoxy group, a cyano group, an amino group, a nitro group, a hydroxyl group, a hydroxyalkyl group, a cycloalkyl group, and a heterocyclic group;

R ⁴ Selected from the group consisting of a hydrogen atom, an alkyl group, a haloalkyl group, an alkoxy group, a haloalkoxy group, a cyano group, an amino group, a nitro group, a hydroxyl group, a hydroxyalkyl group, a cycloalkyl group, and a heterocyclic group;

alternatively, R ³ And R is ⁴ Forming a heterocyclic group with the nitrogen atom to which it is attached, said heterocyclic group optionally being further substituted with a member selected from the group consisting of alkyl, halogen, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, -L ² -C(O)OR ⁹ 、-OR ⁹ and-C (O) R ⁹ Is substituted by one or more substituents;

R ⁵ identical or different and are each independently selected from hydrogen atoms, halogenPlain, alkyl, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclyl, aryl, and heteroaryl;

R ⁶ the same or different and are each independently selected from the group consisting of hydrogen, alkyl, halogen, haloalkyl, amino, hydroxyl, hydroxyalkyl, cycloalkyl, heterocyclyl, aryl, and heteroaryl;

R ⁹ selected from the group consisting of a hydrogen atom, an alkyl group, a haloalkyl group, an alkoxy group, a haloalkoxy group, a cyano group, an amino group, a nitro group, a hydroxyl group, a hydroxyalkyl group, a cycloalkyl group, and a heterocyclic group;

L ² selected from the group consisting of bond, cycloalkyl and alkylene, said cycloalkyl and alkylene being optionally further substituted with one or more substituents selected from the group consisting of alkyl, halogen, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl and heterocyclyl;

r is selected from 0, 1 or 2;

s is selected from 0, 1, 2, 3 or 4; and is also provided with

t is selected from 0, 1, 2, 3, 4 or 5.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein L ¹ Is an alkylene group; preferably C _1-6 An alkylene group; more preferably CH ₂ 。

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein ring a is aryl, preferably phenyl.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound of formula (II) or a tautomer, meso, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein the method comprises the steps of

R ¹ -R ⁶ R, s and t are as defined in formula (I).

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ⁶ Is a hydrogen atom.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound of formula (III) or a tautomer, meso, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ¹ -R ⁵ s and t are as defined in formula (I).

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ³ And R is ⁴ Forming a heterocyclic group with the attached nitrogen atom, said heterocyclic group optionally containing 1 to 2 heteroatoms selected from N, O and S, which are the same or different, in addition to 1 nitrogen atom, and said heterocyclic group optionally being further substituted with a member selected from alkyl, halogen, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclyl, -L ² -C(O)OR ⁹ 、-L ² -OR ⁹ and-L ² -C(O)R ⁹ Is substituted by one or more substituents;

R ⁹ selected from the group consisting of hydrogen, alkyl, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, and heterocyclyl.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ³ And R is ⁴ Forming a heterocyclic group, preferably a 5-6 membered heterocyclic group, with the nitrogen atom to which it is attached; the heterocyclic group optionally contains 1 to 2 identical or different heteroatoms selected from N and O in addition to 1 nitrogen atom, and the heterocyclic group is optionally further substituted by-L ² -C(O)OR ⁹ Substituted;

L ² selected from a bond, cycloalkyl or alkylene, said cycloalkyl or alkylene being optionally further substituted with alkyl;

R ⁹ is a hydrogen atom or an alkyl group.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ³ And R is ⁴ Forming a heterocyclic group selected from morpholinyl, pyrrolidinyl or piperidinyl with the nitrogen atom to which it is attached; said heterocyclyl is optionally further substituted with-L ² -C(O)OR ⁹ Substituted; l (L) ² ，R ⁹ As defined previously.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound of formula (IV) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ⁷ the same or different and are each independently selected from the group consisting of alkyl, halogen, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, and heterocyclyl;

R ⁸ selected from hydrogen atoms, -C (O) OR ⁹ 、-OR ⁹ and-C (O) R ⁹ ；

R ⁹ Is a hydrogen atom or an alkyl group;

u is selected from 0, 1, 2, 3, 4 or 5;

R ¹ 、R ² 、R ⁵ s and t are as defined in formula (I).

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ⁵ Is a hydrogen atom.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound of formula (V) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ^5a and R is ^5b The same or different and are each independently selected from the group consisting of alkyl, halogen, haloalkyl, alkoxy, haloalkoxy, cyano, amino, nitro, hydroxy, hydroxyalkyl, cycloalkyl, and heterocyclyl;

u is selected from 0, 1, 2, 3 or 4;

R ¹ 、R ² 、R ⁷ 、R ⁸ 、L ² u and s are as defined in formula (IV).

In other embodiments of the present disclosure, the compound of formula (I) wherein R ^5a And R is ^5b The same or different, and are each independently alkyl or halogen; preferably, R ^5a And R is ^5b Each independently is C _1-6 Alkyl or halogen.

In other embodiments of the present disclosure, the compound of formula (I) wherein R ⁷ Is a hydrogen atom or an alkyl group, preferably a hydrogen atom or C _1-6 An alkyl group.

In other embodiments of the present disclosure, the compound of formula (I) wherein R ⁸ is-C (O) OH.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein L ² Selected from the group consisting of a bond, a 3-to 8-membered cycloalkyl group, and C _1-6 Alkylene, said 3-to 8-membered cycloalkyl and C _1-6 Alkylene is optionally further selected from C _1-6 Alkyl, halogen and C _1-6 One or more substituents of the haloalkyl group. In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereofWherein L is ² Is a key.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein L ² Is cycloalkyl or alkylene, said cycloalkyl or alkylene being optionally further substituted with alkyl; preferably L ² Selected from CH ₂ 、CH ₂ CH ₂ 、

And->

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ¹ Is alkyl; preferably, R ¹ Is C _1-6 An alkyl group.

In other embodiments of the present disclosure, the compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein R ² Is a hydrogen atom.

Typical compounds of formula (I) or tautomers, racemates, enantiomers, diastereomers, or mixtures thereof, or pharmaceutically acceptable salts thereof, include, but are not limited to:

in another aspect, the present disclosure relates to a compound of formula (IA) or (IVA) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ² 、R ³ 、R ⁴ s is defined as in formula (I); r is R ⁷ 、R ⁸ 、L ² And u is as defined in formula (IV).

Which is an intermediate for preparing a compound represented by the general formula (I) or a tautomer, a racemate, an enantiomer, a diastereomer, or a mixture thereof, or a pharmaceutically acceptable salt thereof.

Typical compounds of formula (IA) include, but are not limited to:

/>

/>

/>

in another aspect, the present disclosure relates to a compound of formula (IB), (IIB), (IIIB) or (VB) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein X is halogen; r is R ¹ 、R ⁵ 、R ⁶ 、L ¹ And t and r are as defined in formula (I).

In another aspect, the present disclosure relates to a compound of formula (VB) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof:

wherein X is halogen; r is R ¹ 、R ^5a And R is ^5b As defined in formula (V).

Typical compounds of formula (IB), (IIB), (IIIB) or (VB) include, but are not limited to:

in another aspect, the present disclosure relates to a method of preparing a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, according to the present disclosure, comprising the steps of:

Nucleophilic substitution reaction is carried out on the compound of the general formula (IA) or salt thereof and the compound of the general formula (IB) under alkaline condition, so as to obtain the compound of the general formula (I);

wherein X is halogen; r is R ¹ -R ⁶ 、L ¹ R, s and t are as defined in formula (I).

Another aspect of the present disclosure relates to a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents, or excipients. The present disclosure also relates to a method of preparing the above composition, comprising mixing a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, with a pharmaceutically acceptable carrier, diluent, or excipient.

The present disclosure further relates to a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, for use as a medicament.

The disclosure further relates to the use of a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, in the preparation of a capsid protein inhibitor.

The disclosure further relates to the use of a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, in the manufacture of a medicament for the prevention and/or treatment of viral infectious diseases. The virus may be hepatitis B virus, influenza virus, herpes virus and HIV, and the disease may be hepatitis B, influenza, herpes and HIV.

The present disclosure further relates to a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, for use as a medicament. The medicament is preferably a medicament for the prophylaxis and/or treatment of viral infectious diseases. The virus may be hepatitis B virus, influenza virus, herpes virus and HIV, and the disease may be hepatitis B, influenza, herpes and HIV.

The present disclosure also relates to compounds of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, for use as a capsid protein inhibitor.

The present disclosure also relates to a method for preventing and/or treating viral infection diseases, which comprises administering to a patient in need thereof a therapeutically effective dose of a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, as a capsid protein inhibitor, or a pharmaceutical composition comprising the same. The virus may be hepatitis B virus, influenza virus, herpes virus and HIV, and the disease may be hepatitis B, influenza, herpes and HIV.

The active compounds may be formulated in a form suitable for administration by any suitable route, preferably in unit dosage form, or in a form whereby the patient may self-administer a single dose. The unit dosage of a compound or composition of the present disclosure may be expressed in the form of a tablet, capsule, cachet, bottled lotion, powder, granule, lozenge, suppository, reconstituted powder or liquid formulation.

The dosage of the compound or composition used in the disclosed methods of treatment will generally vary with the severity of the disease, the weight of the patient, and the relative efficacy of the compound. However, as a general guideline, suitable unit doses may be from 0.1 to 1000mg.

The pharmaceutical compositions of the present disclosure may contain, in addition to the active compound, one or more excipients selected from the following ingredients: fillers (diluents), binders, wetting agents, disintegrants or excipients, and the like. Depending on the method of administration, the compositions may contain from 0.1 to 99% by weight of the active compound.

Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral administration, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Oral compositions may be prepared according to any method known in the art for preparing pharmaceutical compositions, such compositions may contain one or more ingredients selected from the group consisting of: sweeteners, flavoring agents, coloring agents and preservatives to provide a pleasing and palatable pharmaceutical preparation. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients may be inert excipients, granulating agents, disintegrating agents, binding agents, and lubricating agents. These tablets may be uncoated or they may be coated by known techniques to mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.

Oral formulations may also be provided in soft gelatin capsules wherein the active ingredient is mixed with an inert solid diluent or wherein the active ingredient is mixed with a water-soluble carrier or oil vehicle.

Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients are suspensions, dispersants or wetting agents. The aqueous suspension may also contain one or more preservatives, one or more colorants, one or more flavoring agents and one or more sweeteners.

The oil suspensions may be formulated by suspending the active ingredient in a vegetable or mineral oil. The oil suspension may contain a thickener. The above-described sweeteners and flavoring agents may be added to provide a palatable preparation. These compositions can be preserved by the addition of antioxidants.

The pharmaceutical compositions of the present disclosure may also be in the form of an oil-in-water emulsion. The oil phase may be a vegetable oil, or a mineral oil, or a mixture thereof. Suitable emulsifiers may be naturally occurring phospholipids, and emulsions may also contain sweetening, flavoring, preservative and antioxidant agents. Such formulations may also contain a demulcent, a preservative, a colorant and an antioxidant.

The pharmaceutical compositions of the present disclosure may be in the form of sterile injectable aqueous solutions. Acceptable vehicles or solvents that may be used are water, ringer's solution and isotonic sodium chloride solution. The sterile injectable preparation may be a sterile injectable oil-in-water microemulsion in which the active ingredient is dissolved in an oil phase, which injectable solution or microemulsion may be injected into the blood stream of a patient by topical bolus injection. Alternatively, it may be desirable to administer the solutions and microemulsions in a manner that maintains a constant circulating concentration of the compounds of the present disclosure. To maintain this constant concentration, a continuous intravenous drug delivery device may be used. An example of such a device is a Deltec CADD-PLUS. TM.5400 model intravenous pump.

The pharmaceutical compositions of the present disclosure may be in the form of sterile injectable aqueous or oleaginous suspensions for intramuscular and subcutaneous administration. The suspensions may be formulated according to known techniques using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a parenterally-acceptable, nontoxic diluent or solvent. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any blend fixed oil may be used. In addition, fatty acids can also be used to prepare injections.

The compounds of the present disclosure may be administered in the form of suppositories for rectal administration. These pharmaceutical compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid in the rectum and will therefore melt in the rectum to release the drug.

As is well known to those skilled in the art, the amount of drug administered depends on a variety of factors, including, but not limited to, the following: the activity of the specific compound used, the age of the patient, the weight of the patient, the health of the patient, the behavior of the patient, the diet of the patient, the time of administration, the mode of administration, the rate of excretion, the combination of drugs, etc.; in addition, optimal treatment regimens, such as the mode of treatment, daily amounts of a compound of the disclosure, or the type of pharmaceutically acceptable salt, can be verified according to conventional treatment regimens.

The novel structure capsid protein inhibitor shown in the general formula (I) provided by the disclosure comprises a piperazine ureido structure, reduces chiral centers, has obvious inhibition effect on HBV capsid protein normal assembly, and has good drug substitution absorption, high liver exposure, low toxicity and better safety.

Detailed description of the invention

Unless stated to the contrary, the terms used in the specification and claims have the following meanings.

The term "alkyl" refers to a saturated aliphatic hydrocarbon group which is a straight or branched chain group containing from 1 to 20 carbon atoms, preferably an alkyl group containing from 1 to 12 (e.g., 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12) carbon atoms, more preferably an alkyl group containing from 1 to 6 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1-dimethylpropyl, 1, 2-dimethylpropyl, 2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1, 2-trimethylpropyl, 1-dimethylbutyl, 1, 2-dimethylbutyl, 2-dimethylbutyl, 1, 3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2, 3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl 5-methylhexyl, 2, 3-dimethylpentyl, 2, 4-dimethylpentyl, 2-dimethylpentyl, 3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2, 3-dimethylhexyl, 2, 4-dimethylhexyl, 2, 5-dimethylhexyl, 2-dimethylhexyl, 3-dimethylhexyl 4, 4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2-diethylpentyl, n-decyl, 3-diethylhexyl, 2-diethylhexyl, and various branched isomers thereof. More preferred are lower alkyl groups containing 1 to 6 carbon atoms, and non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1-dimethylpropyl, 1, 2-dimethylpropyl, 2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1, 2-trimethylpropyl, 1-dimethylbutyl, 1, 2-dimethylbutyl, 2-dimethylbutyl, 1, 3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2, 3-dimethylbutyl, and the like. The alkyl group may be substituted or unsubstituted, and when substituted, the substituent may be substituted at any available point of attachment, preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl and carboxylate groups.

The term "alkylene" refers to a saturated straight or branched chain aliphatic hydrocarbon group having 2 residues derived from the same carbon atom or two different carbon atoms of the parent alkane, which is a straight or branched chain group containing from 1 to 20 carbon atoms, preferably an alkylene group containing from 1 to 12 (e.g., 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12) carbon atoms, more preferably containing from 1 to 6 carbon atoms. Non-limiting examples of alkylene groups include, but are not limited to, methylene (-CH) ₂ (-), 1-ethylene (-CH (CH) ₃ ) (-), 1, 2-ethylene (-CH) ₂ CH ₂ (-), 1-propylene (-CH (CH) ₂ CH ₃ ) (-), 1, 2-propylene (-CH) ₂ CH(CH ₃ ) (-), 1, 3-propylene (-CH) ₂ CH ₂ CH ₂ (-), 1, 4-butylene (-CH) ₂ CH ₂ CH ₂ CH ₂ (-), etc. Alkylene groups may be substituted or unsubstituted, and when substituted, the substituents may be substituted at any useful point of attachment, preferably independently and optionally selected from H, D, halogen, alkyl, alkoxy, haloalkyl, hydroxy, hydroxyalkyl, cyano, amino, nitro, cycloalkyl,one or more substituents in the heterocyclic, aryl and heteroaryl groups.

The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent, the cycloalkyl ring containing from 3 to 20 carbon atoms, preferably from 3 to 12 carbon atoms (which may be a specific point, or may be a region of optionally two points, such as 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12 ring atoms, 4 to 11 ring atoms, 6 to 12 ring atoms, etc.), more preferably from 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, and the like; polycyclic cycloalkyl groups include spiro, fused and bridged cycloalkyl groups.

The term "spirocycloalkyl" refers to a polycyclic group sharing one carbon atom (referred to as a spiro atom) between 5 to 20 membered monocyclic rings, which may contain one or more double bonds. Preferably 6 to 14 membered, more preferably 7 to 10 membered. The spirocycloalkyl group is classified into a single spirocycloalkyl group, a double spirocycloalkyl group or a multiple spirocycloalkyl group according to the number of common spiro atoms between rings, and preferably a single spirocycloalkyl group and a double spirocycloalkyl group. More preferably 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/5-membered or 5-membered/6-membered monocyclocycloalkyl. Non-limiting examples of spirocycloalkyl groups include:

the term "fused ring alkyl" refers to 5 to 20 membered, all carbon polycyclic groups in which each ring in the system shares an adjacent pair of carbon atoms with the other rings in the system, wherein one or more of the rings may contain one or more double bonds. Preferably 6 to 14 membered, more preferably 7 to 10 membered. The condensed ring alkyl group may be classified as a double ring, triple ring, tetra ring or polycyclic depending on the number of constituent rings, preferably a double ring or triple ring, more preferably a 3-membered/4-membered, 3-membered/5-membered, 3-membered/6-membered, 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/4-membered, 5-membered/5-membered, 5-membered/6-membered, 6-membered/3-membered, 6-membered/4-membered, 6-membered/5-membered and 6-membered/6-membered, and the like. Non-limiting examples of fused ring alkyl groups include:

The term "bridged cycloalkyl" refers to an all-carbon polycyclic group of 5 to 20 members, any two rings sharing two carbon atoms that are not directly attached, which may contain one or more double bonds. Preferably 6 to 14 membered, more preferably 7 to 10 membered. Cycloalkyl groups which may be classified as bicyclic, tricyclic, tetracyclic or polycyclic bridged according to the number of constituent rings are preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of bridged cycloalkyl groups include:

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the cycloalkyl ring includes cycloalkyl (e.g., monocyclic, fused, spiro, and bridged cycloalkyl) fused to an aryl, heteroaryl, or heterocycloalkyl ring as described above, wherein the ring attached to the parent structure is cycloalkyl, non-limiting examples of which include indanyl, tetrahydronaphthyl, benzocycloheptyl, and the like. Cycloalkyl groups may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl and carboxylate groups.

The term "alkoxy" refers to-O- (alkyl) and-O- (unsubstituted cycloalkyl), wherein the alkyl and cycloalkyl are as defined above. Non-limiting examples of alkoxy groups include: methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentoxy, cyclohexyloxy. The alkoxy groups may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl and carboxylate groups.

The term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent containing from 3 to 20 ring atoms in which one or more ring atoms are selected from nitrogen, oxygen or S (O) _m (wherein m is an integer from 0 to 2), but does not include a ring moiety of-O-O-, -O-S-, or-S-S-, and the remaining ring atoms are carbon. Preferably from 3 to 12 (which may be a specific point, or may be an interval of optionally two points, for example 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12 ring atoms, 4 to 11 ring atoms, 6 to 12 ring atoms, etc.) ring atoms, of which 1 to 4 (1, 2, 3 or 4) are heteroatoms; most preferably from 3 to 8 ring atoms, wherein 1 to 3 are heteroatoms; most preferably 3 to 6 ring atoms, of which 1 to 2 are heteroatoms. Non-limiting examples of monocyclic heterocyclyl groups include pyrrolidinyl, imidazolidinyl, tetrahydrofuranyl, tetrahydrothienyl, dihydroimidazolyl, dihydrofuranyl, dihydropyrazolyl, dihydropyrrolyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, pyranyl, and the like, with piperidinyl, piperazinyl, or morpholinyl being preferred. Polycyclic heterocyclyl groups include spiro, fused and bridged heterocyclic groups.

The term "spiroheterocyclyl" refers to a polycyclic heterocyclic group having a single ring of 5 to 20 members sharing one atom (referred to as the spiro atom) between them, wherein one or more of the ring atoms is selected from nitrogen, oxygen or S (O) _m (wherein m is an integer from 0 to 2) and the remaining ring atoms are carbon. Which may contain one or more double bonds. Preferably 6 to 14 membered, more preferably 7 to 10 membered. The spiroheterocyclyl groups are classified into a single spiroheterocyclyl group, a double spiroheterocyclyl group or a multiple spiroheterocyclyl group according to the number of common spiro atoms between rings, and preferably a single spiroheterocyclyl group and a double spiroheterocyclyl group. More preferably 3-membered/6-membered, 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/5-membered or 5-membered/6-membered single spiro heterocyclyl. Non-limiting examples of spiroheterocyclyl groups include:

the term "fused heterocyclyl" refers to a 5 to 20 membered, polycyclic heterocyclic group in which each ring in the system shares an adjacent pair of atoms with the other rings in the system, one or more of which may contain one or more double bonds, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O) _m (wherein m is an integer from 0 to 2) and the remaining ring atoms are carbon. Preferably 6 to 14 membered, more preferably 7 to 10 membered. The number of constituent rings may be classified into a bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic group, preferably a bicyclic or tricyclic, more preferably a 3-membered/4-membered, 3-membered/5-membered, 3-membered/6-membered, 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/4-membered, 5-membered/5-membered, 5-membered/6-membered, 6-membered/3-membered, 6-membered/4-membered, 6-membered/5-membered and 6-membered bicyclic fused heterocyclic group. Non-limiting examples of fused heterocyclyl groups include:

The term "bridged heterocyclyl" refers to a 5 to 14 membered, polycyclic heterocyclic group in which any two rings share two atoms which are not directly attached, which may contain one or more double bonds, wherein one or more ring atoms are selected from nitrogen, oxygen or S (O) _m (wherein m is an integer from 0 to 2) and the remaining ring atoms are carbon. Preferably 6 to 14 membered, more preferably 7 to 10 membered. Heterocyclic groups which may be classified as bicyclic, tricyclic, tetracyclic or polycyclic bridged according to the number of constituent rings are preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of bridged heterocyclyl groups include:

the heterocyclyl ring includes those described above wherein the heterocyclyl (e.g., monocyclic, fused, spiro, and bridged heterocyclyl) is fused to an aryl, heteroaryl, or cycloalkyl ring, wherein the ring attached to the parent structure is a heterocyclyl, non-limiting examples of which include:

the heterocyclic group may be optionally substituted or unsubstituted, and when substituted, the substituent is preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl and carboxylate groups.

The term "aryl" refers to a 6 to 14 membered all-carbon monocyclic or fused polycyclic (i.e., rings sharing adjacent pairs of carbon atoms) group having a conjugated pi-electron system, preferably 6 to 10 membered, such as phenyl and naphthyl. More preferably phenyl. The aryl ring includes aryl fused to heteroaryl, heterocyclyl or cycloalkyl rings as described above, wherein the ring attached to the parent structure is an aryl ring, non-limiting examples of which include:

aryl groups may be substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl, or carboxylate groups.

The term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 (e.g., 1, 2, 3, and 4) heteroatoms, 5 to 14 ring atoms, wherein the heteroatoms are selected from oxygen, sulfur, and nitrogen. Heteroaryl groups are preferably 5 to 10 membered, containing 1 to 3 heteroatoms; more preferably 5 or 6 membered, containing 1 to 2 heteroatoms; preferred are, for example, imidazolyl, furyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, pyrrolyl, tetrazolyl, pyridyl, pyrimidinyl, thiadiazole, pyrazinyl and the like, preferably imidazolyl, tetrazolyl, pyridyl, thienyl, pyrazolyl or pyrimidinyl, thiazolyl; more preferably a pyridyl group. The heteroaryl ring includes those heteroaryl groups described above fused to an aryl, heterocyclyl or cycloalkyl ring, wherein the ring attached to the parent structure is a heteroaryl ring, non-limiting examples of which include:

Heteroaryl groups may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl, and carboxylate groups.

The cycloalkyl, heterocyclyl, aryl and heteroaryl groups described above have 1 residue derived from the removal of one hydrogen atom from the parent carbon atom, or 2 residues derived from the removal of two hydrogen atoms from the same or two different carbon atoms of the parent, 2 residues being "divalent cycloalkyl", "divalent heterocyclyl", "arylene", "heteroarylene".

The term "amino protecting group" is intended to mean an amino group that is protected by an easily removable group in order to keep the amino group unchanged when the reaction is carried out at other positions of the molecule. Non-limiting examples include t-butoxycarbonyl, acetyl, benzyl, allyl, p-methoxybenzyl, and the like. These groups may be optionally substituted with 1 to 3 substituents selected from halogen, alkoxy or nitro. The amino protecting group is preferably p-methoxybenzyl.

The term "ureido" refers to-N-C (O) -N-.

The term "haloalkyl" refers to an alkyl group substituted with one or more halogens, wherein alkyl is as defined above.

The term "haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein the alkoxy group is as defined above.

The term "hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group, wherein alkyl is as defined above.

The term "hydroxy" refers to an-OH group.

The term "halogen" refers to fluorine, chlorine, bromine or iodine.

The term "amino" refers to-NH ₂ 。

The term "cyano" refers to-CN.

The term "nitro" refers to-NO ₂ 。

The term "oxo" refers to = O.

The term "carbonyl" refers to c=o.

The term "carboxy" refers to-C (O) OH.

The term "carboxylate" refers to-C (O) O (alkyl) or-C (O) O (cycloalkyl), wherein alkyl, cycloalkyl are as defined above.

The term "acyl halide" refers to a compound containing a group that is-C (O) -halogen.

"optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs or does not. For example, "a heterocyclic group optionally substituted with an alkyl group" means that an alkyl group may be, but is not necessarily, present, and the description includes cases where the heterocyclic group is substituted with an alkyl group and cases where the heterocyclic group is not substituted with an alkyl group.

"substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in the group are independently substituted with a corresponding number of substituents. It goes without saying that substituents are only in their possible chemical positions, and that the person skilled in the art is able to determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, amino or hydroxyl groups having free hydrogen may be unstable when bound to carbon atoms having unsaturated (e.g., olefinic) bonds.

"pharmaceutical composition" means a mixture comprising one or more of the compounds described herein or a physiologically/pharmaceutically acceptable salt or prodrug thereof, and other chemical components, such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to promote the administration to organisms, facilitate the absorption of active ingredients and thus exert biological activity.

By "pharmaceutically acceptable salts" is meant salts of the compounds of the present disclosure which are safe and effective when used in a mammal, and which possess the desired biological activity.

Methods of synthesizing compounds of the present disclosure

In order to accomplish the purpose of the present disclosure, the present disclosure adopts the following technical scheme:

Scheme one

A process for the preparation of a compound of formula (I) or a tautomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, according to the present disclosure, comprising the steps of:

Scheme II

In some embodiments of the present disclosure, a method of preparing a compound represented by general formula (II) according to the present disclosure, the method comprising the steps of:

carrying out nucleophilic substitution reaction on a compound of a general formula (IA) or salt thereof and a compound of a general formula (IIB) under alkaline conditions to obtain a compound of a general formula (II);

wherein X is halogen; r is R ¹ -R ⁶ R, s and t are as defined in formula (II).

Scheme III

In some embodiments of the present disclosure, a method of preparing a compound represented by general formula (III) according to the present disclosure, the method comprising the steps of:

carrying out nucleophilic substitution reaction on a compound of a general formula (IA) or a salt thereof and a compound of a general formula (IIIB) under alkaline conditions to obtain a compound of a general formula (III);

Wherein X is halogen; r is R ¹ -R ⁵ S and t are as defined in formula (III).

Scheme IV

In some embodiments of the present disclosure, a method of preparing a compound represented by general formula (IV) according to the present disclosure, the method comprising the steps of:

carrying out nucleophilic substitution reaction on a compound of a general formula (IVA) or salt thereof and a compound of a general formula (IIIB) under alkaline conditions to obtain a compound of a general formula (IV);

wherein X is halogen; r is R ¹ 、R ² 、R ⁵ 、R ⁷ 、R ⁸ 、L ² U, s and t are as defined in formula (IV).

Scheme five

In some embodiments of the present disclosure, a method of preparing a compound represented by general formula (V) according to the present disclosure, the method comprising the steps of:

carrying out nucleophilic substitution reaction on a compound of a general formula (IVA) or salt thereof and a compound of a general formula (VB) under alkaline conditions to obtain a compound of a general formula (V);

wherein X is halogen; r is R ¹ 、R ² 、R ^5a 、R ^5b 、R ⁷ 、R ⁸ 、L ² U and s are as defined in formula (V).

In the above preparation scheme, the reagent for providing alkaline conditions comprises organic bases and inorganic bases, wherein the organic bases comprise, but are not limited to, triethylamine, 1M lithium bis (trimethylsilyl) amide tetrahydrofuran solution, N-diisopropylethylamine, N-butyllithium, lithium diisopropylamide, potassium acetate, sodium tert-butoxide or potassium tert-butoxide, and the inorganic bases comprise, but are not limited to, sodium hydride, sodium hydroxide, potassium phosphate, sodium carbonate, sodium bicarbonate, potassium carbonate or cesium carbonate; preferably triethylamine or N, N-diisopropylethylamine;

The above reaction is preferably carried out in a solvent, including but not limited to: acetic acid, methanol, ethanol, toluene, tetrahydrofuran, methylene chloride, petroleum ether, ethyl acetate, N-hexane, dimethyl sulfoxide, 1, 4-dioxane, water, N-dimethylformamide, and mixtures thereof; n, N-dimethylformamide is preferred.

Detailed Description

The present disclosure is further described below in connection with the examples, which are not intended to limit the scope of the present disclosure.

Examples

The structure of the compounds is determined by Nuclear Magnetic Resonance (NMR) or/and Mass Spectrometry (MS). NMR shift (. Delta.) of 10 ^-6 Units of (ppm) are given. NMR was performed using Bruker AVANCE-400 nuclear magnetic resonance apparatus with deuterated dimethyl sulfoxide (DMSO-d) ₆ ) Deuterated chloroform (CDCl) ₃ ) Deuterated methanol (CD) ₃ OD), internal standard is Tetramethylsilane (TMS).

MS was measured using an Agilent 1200/1290DAD-6110/6120 Quadrapol MS liquid chromatography-mass spectrometry (manufacturer: agilent, MS model: 6110/6120 Quadrapol MS), waters ACQuity UPLC-QD/SQD (manufacturer: waters, MS model: waters ACQuity Qda Detector/waters SQ Detector), THERMO Ultimate 3000-Q actual (manufacturer: THERMO, MS model: THERMO Q Exactive).

High Performance Liquid Chromatography (HPLC) analysis used Agilent HPLC1200 DAD, agilent HPLC1200VWD, and Waters HPLC e2695-2489 high pressure liquid chromatography.

Chiral HPLC analysis was determined using an Agilent 1260DAD high performance liquid chromatograph.

The high performance liquid phase was prepared by using a Waters 2545-2767, waters 2767-SQ Detector 2, shimadzu LC-20AP and Gilson GX-281 preparative chromatograph.

Chiral preparation was performed using a Shimadzu LC-20AP preparative chromatograph.

The CombiFlash flash rapid prep instrument used CombiFlash Rf200 (teldyne ISCO).

The thin layer chromatography silica gel plate uses a smoke table yellow sea HSGF254 or Qingdao GF254 silica gel plate, the specification of the silica gel plate used by the Thin Layer Chromatography (TLC) is 0.15 mm-0.2 mm, and the specification of the thin layer chromatography separation and purification product is 0.4 mm-0.5 mm.

The silica gel column chromatography generally uses 200-300 mesh silica gel of yellow sea of the tobacco stand as a carrier.

Average inhibition rate of kinase and IC ₅₀ The values were measured using a NovoStar microplate reader (BMG, germany).

Known starting materials of the present disclosure may be synthesized using or following methods known in the art, or may be purchased from ABCR GmbH & co.kg, acros Organics, aldrich Chemical Company, shaog chemical technology (Accela ChemBio Inc), dary chemicals, and the like.

The examples are not particularly described, and the reaction can be carried out under an argon atmosphere or a nitrogen atmosphere.

An argon or nitrogen atmosphere means that the reactor flask is connected to a balloon of argon or nitrogen of about 1L volume.

The hydrogen atmosphere is defined as the reaction flask being connected to a balloon of hydrogen gas of about 1L volume.

The pressure hydrogenation reaction uses a Parr 3916 model EKX hydrogenometer and a clear blue QL-500 type hydrogen generator or HC2-SS type hydrogenometer.

The hydrogenation reaction is usually vacuumized, filled with hydrogen and repeatedly operated for 3 times.

The microwave reaction used was a CEM Discover-S908860 type microwave reactor.

The examples are not specifically described, and the solution refers to an aqueous solution.

The reaction temperature is room temperature and is 20-30 deg.c without specific explanation in the examples.

The monitoring of the progress of the reaction in the examples employed Thin Layer Chromatography (TLC), the developing reagent used for the reaction, the system of eluent for column chromatography employed for purifying the compound and the developing reagent system of thin layer chromatography included: a: dichloromethane/methanol system, B: n-hexane/ethyl acetate system, C: the volume ratio of the petroleum ether/ethyl acetate system solvent is adjusted according to the polarity of the compound, and can be adjusted by adding a small amount of alkaline or acidic reagents such as triethylamine, acetic acid and the like.

Example 1

(S) -4- (3-fluoro-2-methylphenyl) -6- ((4- (morpholine-4-carbonyl) piperazin-1-yl) methyl) -2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 1

First step

(S) -4- (3-fluoro-2-methylphenyl) -6-methyl-2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 1d

3-fluoro-2-methylbenzaldehyde 1c (20.3 g,146.9538mmol, pickle medical science Co., ltd.) was dissolved in 150mL of isopropyl alcohol, ethyl acetoacetate (19.1 g,146.7634mmol,18.6341 mL), piperidine (1.15 g,13.5060mmol,1.2366 mL), acetic acid (810 mg,13.4883mmol,771.4286 uL) was added, and the reaction was stirred for 4 hours. 2-Thiazolidine hydrochloride 1a (20.0000 g,122.2282mmol, pichia pharmaceutical technology Co., ltd.) was added, triethylamine (14.85 g,146.7536mmol,20.3425 mL) was added dropwise to the reaction mixture for about 20 minutes, and the reaction was stirred overnight and then carried out at 75℃for 7 hours. 200mL of water was added dropwise at room temperature, stirred for 6 hours, filtered, the solid extracted with a mixed solvent of isopropanol and water (V: V=1:1), (15 mL. Times.2), washed with water, dried and purified by chiral preparation (column: CHIRALPAK AS, column size: 5.0cm I.D.x25 cm L,10 μm, mobile phase: meOH=100%, flow rate: 60 mL/min) to give the title compound as 1d (9.87 g), yield: 22.5%.

MS m/z(ESI):360.1[M+1]

Second step

(S) -6- (bromomethyl) -4- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 1e

Compound 1d (1.00000 g,2.7823 mmol) was dissolved in carbon tetrachloride (15 mL), heated to 70deg.C, N-bromosuccinimide (495.2 mg,2.7823 mmol) was added, reacted at 70deg.C for 15min, concentrated under reduced pressure, and purified by silica gel column chromatography with eluent C to give the title compound 1e (950 mg), yield: 77.9%.

MS m/z(ESI):438.0[M+1]

Third step

4- (chlorocarbonyl) piperazine-1-carboxylic acid tert-butyl ester 1g

Piperazine-1-carboxylic acid tert-butyl ester 1f (1.00000 g,5.3691mmol, country reagent) was dissolved in dry 10mL dichloromethane, pyridine (637.0 mg,8.0531mmol,648.0163 uL) was added, trichloromethyl carbonate (1.91190 g,6.4428 mmol) was added at 0deg.C, and the mixture was reacted at 0deg.C for 1 hour and at room temperature for 1 hour. 30mL of methylene chloride was added thereto for dilution, and the mixture was washed with 20mL of 1M diluted hydrochloric acid and dried over anhydrous sodium sulfate. Filtration and concentration under reduced pressure gave 1g (1.19 g) of the crude title compound, which was used in the next reaction without purification.

Fourth step

4- (morpholine-4-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 1h

1g (200 mg, 804.1627. Mu. Mol) of the crude compound and morpholine (140.1 mg,1.6081 mmol) were dissolved in 8mL of anhydrous dichloromethane and stirred at room temperature for 3 hours. 20mL of ethyl acetate was added for dilution, 20mL of 1N diluted hydrochloric acid was added, the mixture was stirred and layered, the organic phase was taken, 10mL of water was washed with water, 10mL of aqueous sodium chloride solution was washed, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the title compound 1h (240 mg), yield: 99.7%.

MS m/z(ESI):244.2[M-55]

Fifth step

Morpholin (piperazin-1-yl) methanone 1i

Compound 1h (240 mg, 801.6969. Mu. Mol) was added to 4mL of a 4M solution of 1, 4-dioxane hydrochloride, stirred for 1 hour, and concentrated under reduced pressure to give the crude title compound 1i (159 mg), which was used in the next reaction without purification.

Sixth step

Compound 1e (200.2 mg, 456.7510. Mu. Mol) was dissolved in 4mL of N, N-dimethylformamide, and morpholine (piperazin-1-yl) methanone (91 mg, 456.7138. Mu. Mol) and triethylamine (138.6 mg,1.3697 mmol) were added. The reaction was carried out at room temperature for 1 hour. Concentrating the reaction solution under reduced pressure, and performing high performance liquid chromatography (column: sharsil-TPrep 21.2×250mm, 5 μm; C18; mobile phase: water (NH) ₄ Ac) and acetonitrile, gradient ratio: 50% -70% acetonitrile) to give the title compound 1 (30 mg), yield: 11.8%.

MS m/z(ESI):557.2[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.60(s,1H),7.82(d,1H),7.42(d,1H),7.27-7.10(m,1H),7.08-7.01(m,1H),6.99-6.91(m,1H),6.62(s,1H),4.07-4.05(m,3H),4.03-3.93(m,1H),3.71-3.70(m,4H),3.41-3.40(m,4H),3.32-3.29(m,4H),2.63-2.61(m,4H),2.56(d,3H),1.13(t,3H)。

Example 2

2- ((S) -1- (4- ((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidine-4-)

Methyl) piperazine-1-carbonyl) pyrrolidin-3-yl) acetic acid 2

First step

(S) -2-pyrrolidin-3-yl) acetic acid 2b

(S) -2- (1- (tert-Butoxycarbonyl) pyrrolidin-3-yl) acetic acid (200 mg, 872.3259. Mu. Mol, pickle medical science and technology Co., ltd.) was added to a 4M solution of 5mL of hydrogen chloride in 1, 4-dioxane and stirred for 1 hour. Concentrated under reduced pressure to give crude title compound 2b (112 mg), which was used in the next reaction without purification.

Second step

(S) -2- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) pyrrolidin-3-yl) acetic acid 2c

1g (215.7 mg, 867.2894. Mu. Mol) of compound 2b (112.0 mg, 867.1644. Mu. Mol) was dissolved in 8mL of anhydrous dichloromethane, triethylamine (219.4 mg,2.1682 mmol) was added with stirring, and the reaction was stirred for 3 hours. 30mL of ethyl acetate was added for dilution, 30mL of 1M diluted hydrochloric acid was added for stirring and delamination, the ethyl acetate phase was taken, washed with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate for filtration and concentration to obtain compound 2c. (296 mg), yield: 99.98%.

Third step

(S) -2- (1-piperazine-1-carbonyl) pyrrolidin-3-yl) acetic acid 2d

Compound 2c (298 mg, 867.0141. Mu. Mol) was dissolved in 5mL of anhydrous dichloromethane, trifluoroacetic acid (5 mL) was slowly added with stirring, and the reaction was stirred for 1 hour. Concentrated under reduced pressure to give the crude title compound 2d (209 mg), which was used in the next reaction without purification.

Fourth step

Methyl) piperazine-1-carbonyl) pyrrolidin-3-yl) acetic acid 2

The crude compound 2d (40 mg, 165.7783. Mu. Mol) was dissolved in 2mL of N, N-dimethylformamide, triethylamine (50.3 mg, 497.0876. Mu. Mol) was added with stirring, and then compound 1e (72.7 mg, 165.8631. Mu. Mol) was added and the reaction was stirred for 1 hour. The reaction solution was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: X-Bridge Prep 30X 150mm; C18; mobile phase: water (0.1% NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 33% -95% acetonitrile) to give the title compound 2. (32 mg), yield: 32.2%.

MS m/z(ESI):599.2[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.60(s,1H),7.82(d,1H),7.41(d,1H),7.08-7.04(m,1H),7.01-6.99(m,1H),6.92-6.88(m,1H),6.02(s,1H),4.10-4.02(m,3H),3.92-3.88(m,1H),3.63-3.58(m,1H),3.48-3.42(m,5H),3.17-3.12(m,1H),2.63-2.62(m,5H),2.54(d,3H),2.47-2.44(m,2H),2.10-2.09(m,2H),1.61-1.56(m,1H),1.14(s,1H),1.12(t,3H)。

Example 3

(S) -1- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) cyclopropane-1-carboxylic acid 3

1- (piperidin-4-yl) cyclopropane-1-carboxylic acid hydrochloride 3b

1- (pyridin-4-yl) cyclopropane-1-carboxylic acid 3a (200 mg,1.23mmol, shanghai Bi-pharmaceutical technologies Co., ltd.) and platinum dioxide (28 mg, 123.31. Mu. Mol, beijing carboline technologies Co., ltd.) were dissolved in 8mL ethanol, concentrated hydrochloric acid (54 mg,1.48 mmol). The mixture was stirred at room temperature under nitrogen ball. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to give the crude title compound 3b (250 mg), which was used in the next reaction without purification.

MS m/z(ESI):170.1[M+1]。

Second step

1- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidin-4-yl) cyclopropane-1-carboxylic acid 3d

Compound 3b (250 mg,1.2155 mmol) and triethylamine (492 mg,4.8621 mmol) were dissolved in 15mL of dichloromethane, compound 3c (302 mg,1.2143 mmol) was added at 0deg.C, stirring was continued for 1 hour, and the reaction was continued at room temperature overnight. The reaction solution was washed with saturated ammonium chloride and sodium chloride solution. The organic phase was concentrated by drying to give compound 3d (420 mg, yield: 90.58%).

MS m/z(ESI):382.1[M+1]。

Third step

1- (1- (piperazine-1-carbonyl) -piperidin-4-yl) cyclopropane-1-carboxylic acid 3e

Compound 3d (180 mg, 471.86. Mu. Mol) was dissolved in 5mL of dichloromethane, trifluoroacetic acid (815 mg,7.15 mmol) was added and stirred at room temperature for 2 hours. The reaction solution was concentrated to give the crude title compound 3e (130 mg), which was used in the next step without purification.

MS m/z(ESI):282.1[M+1]。

Fourth step

Compound 1e (100 mg, 228.6629. Mu. Mol) was dissolved in 5mL of N, N-dimethylformamide, and crude compound 3e (130 mg, 462.0579. Mu. Mol) and triethylamine (92 mg, 909.1807. Mu. Mol) were added. The reaction was carried out at room temperature for 4 hours. The reaction solution was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: X-Bridge Prep 30X 150mm;5 μm; C18; mobile phase: water (0.1% NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 45% -95% acetonitrile) to give the title compound 3 (8 mg, yield: 5.5%).

MS m/z(ESI):639.0[M+1]。

¹ H NMR(400MHz,CDCl ₃ ):δ7.85(s,1H),7.57(s,1H),7.27-7.14(m,2H),6.99-6.97(m,1H),5.99(s,1H),4.83-4.79(m,1H),4.49-4.46(m,2H),4.06-4.04(m,4H),3.78-3.71(m,7H),3.31-3.26(m,2H),2.76-2.74(m,2H),2.42(s,3H),1.67-1.56(m,4H),1.26-1.07(m,3H),1.07-0.81(m,3H)。

Example 4

(S) -4- (3-fluoro-2-methylphenyl) -6- ((4- (4-hydroxypiperidine-1-carbonyl) piperazin-1-yl) methyl) -2- (thiazol-2-)

Phenyl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 4

First step

4- (4-hydroxy-piperidine-1-carbonyl) -piperazine-1-carboxylic acid tert-butyl ester 4b

Compound 1g (285.0 mg,1.1459 mmol) was dissolved in 10mL of anhydrous dichloromethane, and triethylamine (173.9 mg,1.7185 mmol) was added thereto with stirring, followed by stirring for 3 hours. 30mL of ethyl acetate was added, followed by addition of 30mL of 1M diluted hydrochloric acid with stirring, and the layers were separated, the ethyl acetate phase was taken, washed with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude title compound 4b (320 mg), which was used directly in the next reaction without purification.

MS m/z(ESI):258.2[M-55]

Second step

(4-hydroxy-piperidin-1-yl) (piperazin-1-yl) methanone 4c

The crude compound 4b (320 mg,1.0211 mmol) was dissolved in 5mL of anhydrous dichloromethane, 5mL of trifluoroacetic acid was slowly added with stirring, and the reaction was stirred for 1 hour. Concentrated under reduced pressure to give the crude title compound 4c (217 mg), which was used in the next reaction without purification.

Third step

Phenyl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 4

Compound 1e (82.2 mg, 187.5371. Mu. Mol) was dissolved in 1.5mL of N, N-dimethylformamide, and crude compound 4c (40.0 mg, 187.5504. Mu. Mol) and triethylamine (56.9 mg, 562.3119. Mu. Mol) were added thereto and the reaction was stirred for 1 hour. The reaction solution was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: X-Bridge Prep 30X 150mm;5 μm; C18; mobile phase: water (10 mmol NH) ₄ AC) and acetonitrile, gradient ratio: 43% -95% acetonitrile) to afford the title compound 4. (22 mg), yield: 20.6%.

MS m/z(ESI):571.2[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.62(s,1H),7.82(d,1H),7.41(d,1H),7.08-7.04(m,1H),7.01-6.99(m,1H),6.92-6.88(m,1H),6.01(s,1H),4.09-4.02(m,3H),3.93-3.88(m,2H),3.63-3.59(m,2H),3.41-3.35(m,4H),3.02-2.97(m,2H),2.63-2.62(m,3H),2.54(d,3H),1.92-1.90(m,2H),1.31-1.25(m,4H),1.12(t,3H)。

Example 5

(S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) -2-methylpropanoic acid 5

First step

4- (4- (1-ethoxy-2-methyl-1-oxopropan-2-yl) piperidine-1-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 5b

1g (300.0 mg,1.2062 mmol) of ethyl 2-methyl-2- (piperidin-4-yl) propionate hydrochloride (312.8 mg,1.3268mmol, shanghai Haihai Biometrics, inc. Leaching brand) was dissolved in 10mL of anhydrous dichloromethane, and triethylamine (305.1 mg,3.0151 mmol) was added with stirring and the mixture was reacted for 3 hours. 30mL of ethyl acetate was added, followed by 30mL of 1M diluted hydrochloric acid, and the layers were separated, and the ethyl acetate phase was taken, washed with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude title compound 5b (490 mg), which was used in the next reaction without purification.

Second step

2- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidin-4-yl) -2-methylpropanoic acid 5c

Compound 5b (490 mg,1.1907 mmol) was dissolved in 6mL of a mixed solvent of tetrahydrofuran and methanol (V: V=3:1), and 3.5722mL of 1M aqueous solution of lithium hydroxide was added dropwise with stirring, followed by stirring for 2 hours. The reaction solution was concentrated under reduced pressure, 6mL of water and 10mL of diethyl ether were added, 1M diluted hydrochloric acid was added dropwise to pH 4, ethyl acetate was used for extraction (10 mL. Times.3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to give the crude title compound 5c (410 mg), which was used in the next reaction without purification.

MS m/z(ESI):328.2[M-55]

Third step

2-methyl-2- (1-piperazine-1-carbonyl) piperidin-4-yl) propionic acid 5d

The crude compound 5c (410 mg,1.07 mmol) was dissolved in 10mL of anhydrous dichloromethane, 10mL of trifluoroacetic acid was slowly added with stirring, and the reaction was stirred for 1 hour. Concentrated under reduced pressure to give the crude title compound 5d (336 mg), which was used in the next reaction without purification.

Fourth step (S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) -2-methylpropanoic acid 5

Compound 1e (464.0 mg,1.0586 mmol) was dissolved in 7mL of N, N-dimethylformamide, and crude compound 5d (300.0 mg,1.0587 mmol) and triethylamine (321.4 mg,3.1762 mmol) were added and reacted for 1 hour with stirring. The reaction was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: sharsil-T Prep 30 x 150mm;5 μm; C18; mobile phase: water (0.1% TFA) and acetonitrile, gradient ratio: 34% -95% acetonitrile) to give title compound 5 (180 mg), yield: 26.5%.

MS m/z(ESI):641.1[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.62(s,1H),7.97(d,1H),7.75(d,1H),7.22-7.16(m,2H),7.04-7.02(m,1H),6.12(s,1H),5.02(d,1H),4.56(d,1H),4.14-4.01(m,2H),3.86-3.73(m,6H),2.88-2.82(m,2H),2.45(s,3H),1.85-1.82(m,2H),1.69-1.66(m,2H),1.37-1.29(m,5H),1.18(s,6H),1.14(t,3H)。

Example 6

(S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) acetic acid 6

First step

2- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidin-4-yl) acetic acid 6b

1g (100 mg, 402.0813. Mu. Mol) of 2- (piperidin-4-yl) acetic acid 6a (63.3 mg, 442.0913. Mu. Mol, shanghai Shaoshan Yuan reagent technology Co., ltd.) was dissolved in 3mL of anhydrous dichloromethane, and triethylamine (81.4 mg, 804.4321. Mu. Mol) was added with stirring, followed by stirring for 3 hours. 10mL of ethyl acetate was added, followed by 10mL of 1M diluted hydrochloric acid with stirring, and the layers were separated, the ethyl acetate phase was taken, washed with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and compound 6b (140 mg) was concentrated under reduced pressure, yield: 97.96%.

MS m/z(ESI):300.3[M-55]

Second step

2- (1-piperazine-1-carbonyl) piperidin-4-yl) acetic acid 6c

Compound 6b (140 mg, 393.8912. Mu. Mol) was added to 4mL of a 4M solution of 1, 4-dioxane in hydrogen chloride, and the reaction was stirred for 1 hour, concentrated under reduced pressure to give the crude title compound 6c (100 mg), which was used in the next reaction without purification.

Third step (S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) acetic acid 6

Compound 1e (171.7 mg, 391.7290. Mu. Mol) was dissolved in dimethylformamide (3 mL), crude compound 6c (100 mg, 391.6767. Mu. Mol) and triethylamine (118.9 mg,1.1750 mmol) were added and the reaction stirred for 1 hour. The reaction solution was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: boston Prep30 x 150mm;5 μm; C18; mobile phase: water (10 mmol NH) ₄ AC) and acetonitrile, gradient ratio: 33% -95% acetonitrile) to give the title compound 6 (12 mg), yield: 5%.

MS m/z(ESI):613.2[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.62(s,1H),7.82(d,1H),7.42(d,1H),7.08-7.05(m,1H),7.01-7.00(m,1H),6.99-6.91(m,1H),6.02(s,1H),4.10-4.04(m,3H),4.03-3.92(m,1H),3.75-3.72(m,2H),3.39-3.35(m,3H),2.85-2.79(m,2H),2.61-2.56(m,4H),2.55(s,3H),2.31-2.30(m,2H),2.10-2.04(m,1H),1.80-1.77(m,2H),1.32-1.26(m,3H),1.13(t,3H)。

Example 7

(S) -4- (3-fluoro-2-methylphenyl) -6- ((4- (piperidine-1-carbonyl) piperazin-1-yl) methyl) -2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 7

First step

4- (piperidine-1-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 7b

Piperidine 7a (17 mg, 199.6545. Mu. Mol, shanghai Tectam technologies Co., ltd.) was dissolved in 3mL of methylene chloride, triethylamine (30.3 mg, 299.4367. Mu. Mol, 42. Mu.L) was added at 0℃followed by addition of 1g of compound (52.1 mg, 209.4844. Mu. Mol), and the reaction mixture was allowed to warm to room temperature and stirred for 3 hours. The reaction solution was concentrated under reduced pressure to obtain a residue containing the crude title compound 7b, which was directly fed to the next reaction without purification. MS m/z (ESI) 242.2[ M-55].

Second step

Piperazin-1-yl (piperidin-1-yl) methanone hydrochloride 7c

The crude compound 7b (59.4 mg, 199.7363. Mu. Mol) was dissolved in 1.5mL of methylene chloride, and 0.5mL of a 4M 1, 4-dioxane solution of hydrogen chloride was added thereto, followed by stirring for 2 hours. Concentrating under reduced pressure and drying to obtain a residue containing the crude title compound 7c, which was directly fed to the next reaction without purification.

Third step

Compound 1e (70 mg, 159.7032. Mu. Mol) was dissolved in 1.5mL of N, N-dimethylformamide, and the reaction mixture was stirred at room temperature for 16 hours, followed by compound 7c (46.7 mg, 199.7967. Mu. Mol) and triethylamine (48.5 mg, 479.2964. Mu. Mol, 67. Mu.L). After dilution with a small amount of methanol, purification by high performance liquid chromatography (column: sharsil-T Prep 30 x 150mm, 5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 50% -70% acetonitrile) to give the title compound 7 (49 mg), yield: 55.31%.

MS m/z(ESI):555.2[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.64(s,1H),7.82(d,1H),7.41(d,1H),7.09-6.96(m,2H),6.90(t,1H),6.01(s,1H),4.09-3.99(m,3H),3.92-3.87(m,1H),3.86(m,4H),3.24-3.23(m,4H),2.61(t,4H),2.55(d,3H),1.61-1.58(m,6H),1.13(t,3H)。

Example 8

(S) -1- (4- (((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidine-3-carboxylic acid 8

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First step

(S) -4- (3- (methoxycarbonyl) piperidine-1-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 8b

(S) -piperidine-3-carboxylic acid methyl ester hydrochloride 8a (75 mg, 417.4921. Mu. Mol, shanghai Haihong biomedical technology Co., ltd.) was dissolved in 4mL of dichloromethane, triethylamine (105.6 mg,1.0436mmol, 146. Mu.L) and compound 1g (109.1 mg, 438.6707. Mu. Mol) were added in this order at 0℃and the reaction mixture was moved to room temperature and stirred. Dichloromethane was added, stirred with silica gel, and purified by silica gel column chromatography using eluent system C to give the title compound 8b (103 mg) in 69.41% yield.

MS m/z(ESI):356.3[M+1]。

Second step

(S) -1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidine-3-carboxylic acid 8c

Compound 8b (103 mg, 289.7915. Mu. Mol) and lithium hydroxide monohydrate (60.8 mg,1.4489 mmol) were successively dissolved in 6mL of a mixed solution of tetrahydrofuran, methanol and water (V: v=2:1:1), and the reaction was stirred for 16 hours. 1M hydrochloric acid was added dropwise to a solution of pH about 4-5, extracted with ethyl acetate, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and dried to give the title compound 8c (98 mg) as a crude product, which was directly fed to the next reaction without purification.

MS m/z(ESI):342.2[M+1]。

Third step

(S) -1- (piperazine-1-carbonyl) piperidine-3-carboxylic acid hydrochloride 8d

The crude compound 8c (98 mg, 287.0516. Mu. Mol) was dissolved in 1.5mL of dichloromethane and 0.6mL of a 4M solution of 1, 4-dioxane in hydrogen chloride was added and the reaction stirred for 2 hours. Concentrating under reduced pressure, and drying to give the crude title compound 8d (30.4 mg) which was directly added to the next reaction without purification.

MS m/z(ESI):242.2[M+1]。

Fourth step

Compound 1e (40 mg, 159.7032. Mu. Mol) was dissolved in 1.0mL of N, N-dimethylformamide, and crude compound 8d (30.4 mg, 109.4521. Mu. Mol) and triethylamine (27.7 mg, 273.7425. Mu. Mol, 38. Mu.L) were added thereto and the reaction was stirred for 16 hours. After dilution with a small amount of methanol, purification by high performance liquid chromatography (column: sharsil-T Prep 30 x 150mm, 5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 30% -50% acetonitrile) to give the title compound 8 (26 mg), yield: 47.59%.

MS m/z(ESI):599.1[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.61(s,1H),7.82(d,1H),7.41(d,1H),7.09-7.04(m,1H),7.01-6.99(m,1H),6.90(t,1H),6.01(s,1H),4.10-3.99(m,3H),3.92-3.88(m,1H),3.71(d,2H),3.50(d,2H),3.39(brs,4H),3.19-2.97(m,2H),2.61(m,3H),2.54(s,3H),2.03-1.55(m,4H),1.12(t,3H)。

Example 9

(S) -3- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) propionic acid 9

First step

4- (4- (3-methoxy-3-oxopropyl) piperidine-1-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 9b

1g (100.0 mg, 402.0813. Mu. Mol) of methyl (3-piperidin-4-yl) propionate 9a (75.7 mg, 442.0797. Mu. Mol, shanghai Shaoshan Yuan reagent technology Co., ltd.) was dissolved in 3mL of anhydrous methylene chloride, and triethylamine (81.4 mg, 804.4273. Mu. Mol) was added with stirring, followed by stirring for 3 hours. 10mL of ethyl acetate was added, followed by 10mL of 1M diluted hydrochloric acid with stirring, and the layers were separated, and the ethyl acetate phase was washed successively with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the title compound 9b (150 mg), yield: 97.28%.

Second step

3- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidin-4-yl) propionic acid 9c

Compound 9b (150 mg,391.1536 μmol) was dissolved in 2mL of a mixed solution of tetrahydrofuran and methanol (V: v=3:1), and the reaction was stirred, lithium hydroxide/water (1 m,1.1724 mL) was added dropwise, and the reaction was stirred for 2 hours. The reaction solution was concentrated under reduced pressure, 3mL of water and 5mL of diethyl ether were sequentially added, the mixture was extracted with 1M diluted hydrochloric acid to pH 4, ethyl acetate (6 ml×3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the title compound 9c (140 mg), yield: 96.88%.

MS m/z(ESI):314.2[M-55]

Third step

3- (1-piperazine-1-carbonyl) piperidin-4-yl) propionic acid 9d

Compound 9c (150 mg, 406.0039. Mu. Mol) was added to 4mL of a 4M solution of 1, 4-dioxane in hydrogen chloride, and the reaction was stirred for 1 hour. The reaction solution was concentrated under reduced pressure to give the crude title compound 9d (109 mg), which was used in the next reaction without purification.

MS m/z(ESI):270.1[M+1]

Fourth step (S) -3- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidin-4-yl) propionic acid 9

Compound 1e (171.7 mg, 391.7518. Mu. Mol) was dissolved in 3mL of N, N-dimethylformamide, and crude compound 9d (105.5 mg, 391.6995. Mu. Mol) and triethylamine (118.9 mg,1.1750 mmol) were added. The reaction was carried out at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: sharsil-TPrep 30 x 150mm;5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 40% -55% acetonitrile) to give the title compound 9 (30 mg), yield: 12.22%.

MS m/z(ESI):627.0[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.64(s,1H),7.83(d,1H),7.42(d,1H),7.09-7.07(m,1H),7.02-7.00(m,1H),6.91-6.89(m,1H),6.02(s,1H),4.10-4.03(m,3H),4.03-3.92(m,1H),3.88-3.72(m,2H),3.38(s,3H),2.73-2.77(m,2H),2.62(s,4H),2.55(s,3H),2.41-2.37(m,2H),2.09-2.04(m,2H),1.73-1.70(m,2H),1.63-1.61(m,2H),1.15-1.14(m,2H),1.12(t,3H)。

Example 10

(S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) -4-methylpiperidin-4-yl) acetic acid 10

First step

2- (4-Methylpiperidin-4-yl) acetic acid 10b

2- (1- (t-Butoxycarbonyl) -4-methylpiperidin-4-yl) acetic acid 10a (300mg,1.1658mmol,Ark pharm reagent) was added to 6mL of a 1, 4-dioxane solution of 4M hydrogen chloride, and the reaction was stirred for 1 hour, concentrated under reduced pressure to give the crude title compound 10b (183 mg), which was used in the next reaction without purification. MS m/z (ESI): 158.1[ M+1]

Second step

2- (1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) -4-methylpiperidin-4-yl) acetic acid 10c

1g (285.0 mg,1.1459 mmol) of compound 10b (180.2 mg,1.1462 mmol) was dissolved in anhydrous dichloromethane (10 mL) and triethylamine (289.9 mg,2.8649 mmol) was added with stirring and stirred for 3 hours. 30mL of ethyl acetate was added to dilute, followed by 30mL of 1M diluted hydrochloric acid, stirring, and the layers were separated, the ethyl acetate phase was taken, washed with 10mL of water, 10mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the title compound 10c (423 mg), yield: 99.9%.

Third step

2- (4-methyl-1- (piperazine-1-carbonyl) piperidin-4-yl) acetic acid 10d

Compound 10c (423 mg,1.1449 mmol) was dissolved in 10mL of anhydrous dichloromethane, 10mL of trifluoroacetic acid was slowly added with stirring, and the reaction was stirred for 1 hour. Concentrated under reduced pressure to give the crude title compound 10d (308 mg), which was used in the next reaction without purification.

Fourth step (S) -2- (1- (4- ((5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) -4-methylpiperidin-4-yl) acetic acid 10

Compound 1e (146.5 mg, 334.2359. Mu. Mol) was dissolved in 2mL of N, N-dimethylformamide, and crude compound 10d (90.0 mg, 334.1513. Mu. Mol) and triethylamine (101.4 mg,1.0021 mmol) were added thereto and reacted under stirring for 1 hour. The reaction was concentrated under reduced pressure and the residue was purified by high performance liquid chromatography (column: sunfire Prep 19 x 150mm;5 μm; C18; mobile phase: water (0.1% TFA) and acetonitrile, gradient ratio: 25% -95% acetonitrile) to give the title compound 10. (25 mg), yield: 11.94%.

MS m/z(ESI):627.3[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.60(s,1H),7.86(d,1H),7.59(d,1H),7.18-7.15(m,2H),7.00-6.96(m,1H),6.00(s,1H),4.84-4.80(m,1H),4.50-4.46(m,1H),4.06-3.99(m,3H),3.71(s,4H),3.38-3.34(m,2H),3.26-3.16(m,2H),2.42(s,3H),2.32(s,2H),1.65-1.55(m,2H),1.52-1.36(m,2H),1.34-1.26(m,3H),1.14(s,3H),1.10(t,3H)。

Example 11

1- ((R) -4- (((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl)

Methyl) -2-methylpiperazine-1-carbonyl) piperidine-4-carboxylic acid 11

First step

(R) -4- (chlorocarbonyl) -3-methylpiperazine-1-carboxylic acid tert-butyl ester 11b

(3R) -3-methylpiperazine-1-carboxylic acid tert-butyl ester 11a (150 mg, 748.9619. Mu. Mol, shanghai Bi-pharmaceutical technologies Co., ltd.) and pyridine (90.6 mg,1.1454mmol, 92. Mu.L) were dissolved in methylene chloride (2.5 mL), cooled to 0℃under nitrogen, and a solution of trichloromethyl carbonate (266.8 mg, 899.0784. Mu. Mol) in methylene chloride (2.5 mL) was slowly added thereto, and stirred at 0℃for 1 hour and then cooled to room temperature and stirred for 1 hour. To the reaction mixture was added 1M hydrochloric acid, followed by addition of dichloromethane and water, and the organic phase was separated, washed with a saturated sodium chloride solution and dried over anhydrous sodium sulfate. Filtration, concentration under reduced pressure and drying gave residue 11b (192 mg) containing the title compound, which was directly fed to the next reaction without purification.

Second step

(R) -4- (4- (methoxycarbonyl) piperidine-1-carbonyl) -3-methylpiperazine-1-carboxylic acid tert-butyl ester 11c

Piperidine-4-carboxylic acid methyl ester (100 mg, 698.4064. Mu. Mol, 94. Mu.L, shanghai Kaisha far reagent technologies Co., ltd.) was dissolved in methylene chloride (7.5 mL), triethylamine (141.3 mg,1.3964mmol, 195. Mu.L), compound 11b (192 mg, 733.4459. Mu. Mol) was added in this order at 0℃and the reaction mixture was stirred at room temperature for 2 hours. Silica gel was added, concentrated under reduced pressure, and the residue was purified by silica gel column chromatography with eluent system C to give the title compound 11C (233 mg), yield: 90.3%.

MS m/z(ESI):314.1[M-55]。

Third step

(R) -1- (4- (tert-Butoxycarbonyl) -2-methylpiperazine-1-carbonyl) piperidine-4-carboxylic acid 11d

Compound 11c (233 mg, 630.6587. Mu. Mol) and lithium hydroxide monohydrate (105.9 mg,2.5236 mmol) were successively dissolved in 6mL of a mixed solution of tetrahydrofuran, methanol and water (V: v=2:1:1), and reacted for 16 hours with stirring. 1M hydrochloric acid was added dropwise to a solution pH of about 4-5, extracted with ethyl acetate, washed with a saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and dried to give crude compound 11d (215 mg), which was directly fed to the next reaction without purification.

MS m/z(ESI):300.1[M-55]。

Fourth step

(R) -1- (2-methylpiperazine-1-carbonyl) piperidine-4-carboxylic acid hydrochloride 11e

Compound 11d (215 mg, 604.9045. Mu. Mol) was dissolved in 1.5mL of methylene chloride, and 1.2mL of a 4M 1, 4-dioxane solution of hydrogen chloride was added thereto, followed by stirring and reaction for 2 hours. Concentrating under reduced pressure, and drying to obtain crude compound 11e (40 mg), which is directly fed to the next reaction without purification.

Fifth step

1- ((R) -4- (((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -2-methylpiperazine-1-carbonyl) piperidine-4-carboxylic acid 11

Compound 11e (40.0 mg, 137.0926. Mu. Mol) was dissolved in 1mL of N, N-dimethylformamide, triethylamine (34.6 mg, 341.9310. Mu. Mol, 48. Mu.L) and compound 1e (50 mg, 114.0737. Mu. Mol) were added in this order at 0℃and stirred at room temperature for 16 hours. The reaction mixture was diluted with a small amount of methanol, filtered through a filter membrane and purified by high performance liquid chromatography (column: sharsil-T Prep 30 x 150mm;5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 30% -50% acetonitrile) to give the title compound 11 (64 mg), yield: 91.57%.

MS m/z(ESI):613.0[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.70(s,1H),7.80(d,1H),7.41(d,1H),7.07(q,1H),7.00-6.96(m,1H),6.90(t,1H),6.01(s,1H),4.11-3.99(m,4H),3.86-3.82(m,1H),3.66-3.64(m,2H),3.40-3.39(m,2H),2.88(q,2H),2.68-2.61(m,3H),2.55(s,3H),2.51-2.34(m,1H),1.97-1.94(m,4H),1.75-1.68(m,1H),1.45(d,3H),1.12(t,3H).

Example 12

(R) -1- (4- (((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) piperazine-1-carbonyl) piperidine-3-carboxylic acid 12

First step

(R) -4- (3- (methoxycarbonyl) piperidine-1-carbonyl) piperazine-1-carboxylic acid tert-butyl ester 12b

(R) -piperidine-3-carboxylic acid methyl ester hydrochloride 12a (80 mg, 445.3250. Mu. Mol, nanjing medical science Co., ltd.) was dissolved in 4mL of methylene chloride, and triethylamine (112.7 mg,1.1137mmol, 155. Mu.L) and compound 1g (116.3 mg, 467.6206. Mu. Mol) were added in this order at 0℃to stir the reaction mixture at room temperature. LC-MS monitoring showed complete reaction of the starting material, addition of dichloromethane, stirring with silica gel, concentration under reduced pressure, and purification of the residue by silica gel column chromatography using eluent system C gave title compound 12b (97 mg) in 61.28% yield.

MS m/z(ESI):356.2[M+1]。

Second step

(R) -1- (4- (tert-Butoxycarbonyl) piperazine-1-carbonyl) piperidine-3-carboxylic acid 12c

Compound 12b (97 mg,272.9103 μmol) and lithium hydroxide monohydrate (57.3 mg,1.3655 mmol) were dissolved in 4mL of a mixed solution of tetrahydrofuran, methanol and water (V: v=2:1:1) in this order, and the reaction was stirred for 16 hours. 1M hydrochloric acid was added dropwise to the solution until the pH was about 4-5, extracted with ethyl acetate, washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. Filtration, concentration under reduced pressure and drying gave crude compound 12c (79 mg) which was directly fed to the next reaction without purification.

MS m/z(ESI):342.2[M+1]。

Third step

(R) -1- (piperazine-1-carbonyl) piperidine-3-carboxylic acid hydrochloride 12d

Compound 12c (79 mg, 231.3990. Mu. Mol) was dissolved in 1mL of methylene chloride, and 0.46mL of a 4M solution of 1, 4-dioxane of hydrogen chloride was added thereto, followed by stirring for reaction for 2 hours. Concentrating under reduced pressure, and drying to obtain crude compound 12d (61.8 mg), which is directly fed to the next reaction without purification.

MS m/z(ESI):242.2[M+1]。

Fourth step

Will be converted intoCompound 1e (75 mg, 171.1105. Mu. Mol) was dissolved in 1.8mL of N, N-dimethylformamide, and Compound 12d (61.8 mg, 222.5046. Mu. Mol) and triethylamine (52.0 mg, 513.8848. Mu. Mol, 72. Mu.L) were added thereto and the reaction was stirred for 16 hours. After dilution with a small amount of methanol, purification by high performance liquid chromatography (column: sharsil-T Prep 30 x 150mm, 5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 30% -50% acetonitrile) to give the title compound 12 (73 mg), yield: 71.26%.

MS m/z(ESI):599.3[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.60(s,1H),7.81(d,1H),7.40(d,1H),7.09-7.04(m,1H),7.00-6.96(m,1H),6.90(t,1H),6.01(s,1H),4.09-3.87(m,3H),3.39(brs,4H),3.02(m,3H),2.77-2.61(m,7H),2.54(s,3H),2.03-1.68(m,4H),1.12(t,3H)。

Example 13

2- (1- ((R) -4- ((S) -5- (ethoxycarbonyl) -6- (3-fluoro-2-methylphenyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidine-4-)

Methyl) -2-methylpiperazine-1-carbonyl) piperidin-4-yl) acetic acid 13

First step

(R) -4-chlorocarbonyl-3-methylpiperazine-1-carboxylic acid tert-butyl ester 13b

Tert-butyl (R) 3-methylpiperidine-1-carboxylate (1 g,4.9931mmol, pichia pharmaceutical technology Co., ltd.) was dissolved in 10mL of dried dichloromethane, pyridine (592.4 mg,7.4893mmol,602.6450 uL) was added, trichloromethyl carbonate (1.778 g,5.9916 mmol) was added at 0℃and reacted for 1 hour at 0℃with stirring. 30mL of methylene chloride was added, and the mixture was washed with 20mL of 1M diluted hydrochloric acid and dried over anhydrous sodium sulfate. Filtration and concentration under reduced pressure gave the title compound 13b (1.3 g), yield: 99%.

Second step

(R) -2- (1- (4- (tert-Butoxycarbonyl) -2-methylpiperazine-1-carbonyl) piperidin-4-yl) acetic acid 13c

Compound 13b (1.4160 g,5.3895 mmol), 2- (4-piperidinyl) acetic acid (848.9 mg,5.9288mmol, shanghai Shaoshan reagent technologies Co., ltd.) was dissolved in 40mL of dry dichloromethane, and triethylamine (1.09070 g,10.7787 mmol) was added with stirring and the reaction was stirred for 3 hours. 30mL of ethyl acetate was added, followed by 30mL of 1M diluted hydrochloric acid, stirring, delamination, ethyl acetate phase taking, washing with 10mL of water, 10mL of saturated sodium chloride solution, drying over anhydrous sodium sulfate, filtration, and concentration under reduced pressure gave the title compound 13c (1.78 g), yield: 89.39%.

Third step

(R) -2- (1- (2-methylpiperazine-1-carbonyl) piperidin-4-yl) acetic acid 13d

Compound 13c (100 mg, 270.6693. Mu. Mol) was dissolved in 5mL of anhydrous dichloromethane, and 5mL of trifluoroacetic acid was slowly added with stirring, followed by stirring for 1 hour. Concentrated under reduced pressure to give the crude title compound 13d (72 mg), which was used in the next reaction without purification.

Fourth step

Methyl) -2-methylpiperazine-1-carbonyl) piperidin-4-yl) acetic acid 13

(Compound 1e (118.6 mg, 270.5828. Mu. Mol) was dissolved in 2 mM N, N-dimethylformamide, and Compound 13d (72.90 mg, 270.6625. Mu. Mol) and triethylamine (82.2 mg, 812.3332. Mu. Mol) were added thereto and reacted for 1 hour with stirring, the reaction mixture was concentrated under reduced pressure and purified by high performance liquid chromatography (column: sharsil-Tprep 21.2X 250mm, 5. Mu. M, C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 50% -70% acetonitrile) to give the title compound 13 (25 mg), yield: 14.74%.

MS m/z(ESI):627.1[M+1]

¹ H NMR(400MHz,CDCl ₃ ):δ9.60(s,1H),7.92(d,1H),7.67(d,1H),7.21-7.18(m,1H),7.13-7.11(m,1H),7.05-7.02(m,1H),6.06(s,1H),4.80-4.77(m,1H),4.61-4.57(m,1H),4.15-4.02(m,3H),3.64-3.63(m,1H),2.93-2.87(m,2H),2.67-2.55(m,1H),2.46(s,3H),2.36-2.35(m,1H),2.34-2.31(m,2H),2.12-2.05(m,2H),1.85-1.82(m,2H),1.55-1.54(m,3H),1.35-1.25(m,5H),1.14(t,3H),0.91-0.90(m,1H)。

Example 14

2- (1- ((R) -4- (((R) -5- (ethoxycarbonyl) -2- (thiazol-2-yl) -6- (2, 3, 4-trifluorophenyl) -3, 6-dihydropyrimidine-4-)

Methyl) -2-methylpiperazine-1-carbonyl) piperidin-4-yl) acetic acid 14

First step

(R) -6-methyl-2- (thiazol-2-yl) -4- (2, 3, 4-trifluorophenyl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 14d

2,3, 4-trifluorobenzaldehyde 14c (5.626 g,35.1408mmol, hadamard) was dissolved in 50mL of isopropanol, and compound 1b (4.573 g,35.1402mmol,4.4617 mL), piperidine (287.5 mg,3.3765 mmol) and acetic acid (201.9 mg,3.3621mmol,192.2857 uL) were added and reacted with stirring for 4 hours. Compound 1a (5 g,30.5570mmol, pickle medical science Co., ltd.) was added dropwise triethylamine (3.7 g,36.5649mmol,5.0685 mL) to the reaction mixture for about 20 minutes, and the reaction was stirred overnight and allowed to react at 85℃for 8 hours. After cooling to room temperature, 50mL of water was added dropwise, stirring was completed for 6 hours, and the solid was extracted with a mixed solvent of isopropyl alcohol and water (V: v=1:1), (15 ml×2), washed with water, dried, and purified by chiral preparation (column: CHIRALPAK AS, column size: 5.0cm I.D.x25 cm L,10 μm, mobile phase: meoh=100%, flow rate: 60 mL/min) to give the title compound 14d (6.1 g), yield: 52.3444%.

MS m/z(ESI):382.0[M+1]。

Second step

(R) -6- (bromomethyl) -2- (thiazol-2-yl) -4- (2, 3, 4-trifluorophenyl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 14e

Compound 14d (337.5 mg, 884.9636. Mu. Mol) was dissolved in carbon tetrachloride (7 mL), heated to 70℃and N-bromosuccinimide (157.5 mg, 884.9115. Mu. Mol) was added thereto, reacted at 70℃for 15 minutes, concentrated under reduced pressure, and purified by silica gel column chromatography with eluent C to give the title compound 14e (284 mg), yield: 69.724%. MS m/z (ESI) 460.0[ M+1],462.0[ M+1].

Third step

2- (1- ((R) -4- (((R) -5- (ethoxycarbonyl) -2- (thiazol-2-yl) -6- (2, 3, 4-trifluorophenyl) -3, 6-dihydropyrimidin-4-yl) methyl) -2-methylpiperazin-1-carbonyl) piperidin-4-yl) acetic acid 14

Compound 14e (284 mg, 617.0322. Mu. Mol) was dissolved in 6mL dimethylformamide, and Compound 13d (249.3 mg, 925.5990. Mu. Mol) and N, N diisopropylethylamine (199.4 mg,1.5428 mmol) were added. The reaction was carried out at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure and purified by high performance liquid chromatography (column: sharsil-TPrep 21.2 x 250mm;5 μm; C18; mobile phase: water (10 mmol NH) ₄ HCO ₃ ) And acetonitrile, gradient ratio: 40% -95% acetonitrile) to afford title compound 14 (266 mg), yield: 66.4559%.

MS m/z(ESI):649.2[M+1]。

¹ H NMR(400MHz,CDCl ₃ ):δ9.82(s,1H),7.85(d,1H),7.46(d,1H),7.05-7.03(m,1H),6.92-6.88(m,1H),6.03(s,1H),4.10-4.04(m,2H),3.93-3.89(m,2H),3.78-3.69(m,3H),3.39-3.38(m,2H),2.82-2.79(m,3H),2.68-2.56(m,5H),2.37-2.31(m,3H),2.10-2.03(m,1H),1.79-1.76(m,2H),1.45(d,3H),1.18(t,3H)。

Example 15

1- (1- ((R) -4- (((R) -6- (2-chloro-3-fluorophenyl) -5- (ethoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -2-methylpiperazin-1-carbonyl) piperidin-4-yl) cyclopropanecarboxylic acid 15

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First step

(R) -4- (2-chloro-3-fluorophenyl) -6-methyl-2-thiazol-2-yl-1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 15d

Compound 2-chloro-3-fluorobenzaldehyde 15c (3.0 g,18.9206mmol, pichia pharmaceutical technology Co., ltd.) was dissolved in isopropanol (22.5 mL), and compound 1b (2.5 g,19.2099mmol,2.4390mL, guog reagent), piperidine (150 mg,1.7617 mmol), acetic acid (106 mg,1.7651mmol,100.9524 uL) was added at room temperature and reacted at room temperature for 4 hours. Compound 1a (2.6 g,15.8897mmol, hcl, distal to shao) was added and triethylamine (1.94 g,19.1719mmol,2.6575 ml) was added dropwise for about 20 minutes, and the reaction was completed at room temperature overnight. After reaction at 75 ℃ for 7 hours, cooling to room temperature, dropwise adding 200mL of water, stirring for 6 hours, filtering, extracting the solid with a mixed solvent of isopropyl alcohol and water (V: v=1:1), washing with water, (15 ml×2), drying, purifying with chiral preparation (chromatographic column: CHIRALPAK AS, column size: 5.0cm I.D.x25 cm L,10 μm, mobile phase: meoh=100%, flow rate: 60 mL/min), to give the title compound 15d (2.02 g), yield: 33.5%.

MS m/z(ESI):380.1[M+1]。

Second step

(R) -6- (bromomethyl) -4- (2-chloro-3-fluorophenyl) -2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylic acid ethyl ester 15e

Compound 15d (470 mg,1.2374 mmol) was dissolved in carbon tetrachloride (5 mL), N-bromosuccinimide (495.2 mg,2.7823 mmol) was added, reacted at room temperature for 2 hours, concentrated under reduced pressure, and purified by silica gel column chromatography with eluent C to give the title compound 15e (340 mg), yield: 59.9%.

MS m/z(ESI):458.8[M+1]。

Third step

15g of 1- (piperidinyl-4-yl) cyclopropanecarboxylic acid

1- (pyridin-4-yl) cyclopropanecarboxylic acid 15f (1 g,6.1285mmol, shao Yuan), platinum dioxide (150 mg, 660.5689. Mu. Mol) and hydrochloric acid (corrosive) (Yi Zhu-toxin-3) (230 mg,6.3081 mmol) were dissolved in ethanol (8 mL.) and replaced with hydrogen 3 times, and stirred overnight at 30 ℃. The reaction solution was filtered, and the filtrate was concentrated to give the title compound 15g (1.2 g), yield: 95.2%.

MS m/z(ESI):170.1[M+1]。

Fourth step

(R) -1- (4- (4- (tert-butoxycarbonyl) -2-methylpiperazine-1-carbonyl) piperidin-4-yl) cyclopropanecarboxylic acid 15h

Compound 15g (390 mg,1.9059mmol, HCl) and triethylamine (578 mg,5.7120 mmol) are dissolved in dichloromethane (15 mL). Compound 11b (500 mg,1.9031 mmol) was added at 0deg.C. The reaction was stirred at room temperature overnight. The reaction solution was diluted with dichloromethane, 1N hydrochloric acid washed, and concentrated by organic phase drying to give the title compound (740 mg), yield: 98.3%.

MS m/z(ESI):396.1[M+1]。

Fifth step

(R) -2- (1- (2-methylpiperazine-1-carbonyl) piperidin-4-yl) cyclopropanecarboxylic acid 15i

Compound 15h (740 mg,1.8711 mmol) was added to 5mL of 4M 1, 4-dioxane solution of hydrochloric acid, stirred for 1 hour, and concentrated under reduced pressure to give the crude title compound 15i (620 mg) which was used in the next reaction without purification.

Sixth step

Compound 15e (70 mg, 152.9239. Mu. Mol) was dissolved in 3mL of dimethylformamide, followed by N, N-diisopropylethylamine (60 mg, 464.2418. Mu. Mol) and compound 15i (76 mg, 229.0279. Mu. Mol, HCl). The reaction was carried out at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure and purified by high performance liquid chromatography (column: gemini Prep 21.2 x 250mm;5 μm; C18; mobile phase: water (10 mmol CH) ₃ COONH ₄ ) And acetonitrile, gradient ratio: 40% -95% acetonitrile) to give the title compound 15 (10 mg), yield: 9.7%.

MS m/z(ESI):673.1[M+1]。

¹ H NMR(400MHz,CDCl ₃ ):δ9.69(s,1H),7.76-7.75(m,1H),7.37-7.36(m,1H),7.19-7.05(m,2H),6.98-6.96(m,1H),6.20(s,1H),3.97-3.90(m,4H),3.76-3.72(m,1H),3.67-3.65(m,2H),3.32-3.30(m,2H),2.66-2.56(m,5H),2.38-2.31(m,1H),1.58-1.56(m,4H),1.39-1.37(m,4H),1.18-1.06(m,4H),1.04-1.03(m,3H)。

Example 16

2- (1- ((R) -4- (((R) -6- (2-chloro-3-fluorophenyl) -5- (ethoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -2-methylpiperazin-1-carbonyl) piperidin-4-yl) acetic acid 16

First step

Compound 15e (200 mg, 436.9257. Mu. Mol) was dissolved in 4mL of dimethylformamide, followed by N, N-diisopropylethylamine (170 mg,1.3154 mmol) and compound 13d (180 mg, 668.3025. Mu. Mol). The reaction was carried out at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure and purified by high performance liquid chromatography (column: gemini Prep 21.2 x 250mm;5 μm; C18; mobile phase: water (10 mmol CH) ₃ COONH ₄ ) And acetonitrile, gradient ratio: 50% -95% acetonitrile) to afford title compound 16 (118 mg), yield: 41.7%.

MS m/z(ESI):647.3[M+1]。

¹ H NMR(400MHz,CDCl ₃ ):δ9.74(s,1H),7.82(d,1H),7.43(d,1H),7.26-7.12(m,2H),7.05-7.03(m,1H),6.27(s,1H),4.04-3.96(m,4H),3.83-3.79(m,1H),3.69-3.63(m,2H),3.39-3.37(m,2H),2.82-2.63(m,6H),2.36-2.30(m,4H),2.17-1.91(m,2H),1.79-1.76(m,2H),1.45-1.44(m,3H),1.13-1.09(m,3H)。

Biological evaluation

Test example 1, in vitro anti-HBV Activity test of the public Compounds (intracellular HBV DNA quantitative analysis)

1. Experimental material and instrument

1.QIAamp 96DNAQIAcube HT Kit(Qiagen)

2.QIAcube HT plasticware(Qiagen)

3. Hepatitis B virus nucleic acid quantitative determination reagent box (Taipu organism)

DNA extraction apparatus (QIAcube) (Qiagen)

5.QuantStudio 6Fiex(ABI,ThermFisher)

6. Enzyme label instrument (BMG)

HepG2.2.15 cells (Shanghai Rei deer Biotechnology Co., ltd.)

2. Experimental procedure

HepG2.2.15 cells are stably expressed cell lines integrating HBV genomes and can be secreted extracellularly by replication, transcription, translation, and packaging into viral particles with HBV DNA. The quantitative PCR method is adopted to quantitatively analyze HBV DNA generated by in vitro proliferation of HepG2.2.15, and the activity of the compound disclosed by the invention for inhibiting HBV DNA replication through the assembly inhibition of HBV capsid protein is screened.

HepG2.2.15 cells were cultured in DMEM/high glucose medium (10% FBS, 400. Mu.g/ml G418) and passaged every 3 days. Cell suspensions were prepared on the day of the experiment in fresh cell culture medium and incubated in 40,000 cells/well 96-well plates (Corning, # 3599), 5% carbon dioxide, at 37 ℃. The next day, compounds were first dissolved in pure DMSO at 20mM, then prepared with DMSO at the first concentration of 2mM, and diluted sequentially 4-fold to 8 concentrations, and 90 μl DMSO was added to wells where controls were placed. Diluted 200-fold with DMEM/high glucose medium. The cell culture plate inoculated on the first day was taken out, the medium in the well plate was sucked out by a negative pressure suction device, and the prepared medium containing the compound of each concentration was added to each well, and cultured at 200. Mu.l/well for 72 hours at 37 ℃. The fifth day, the cells were changed with fresh medium containing the same compound, the method was the same as the second day, and cultured at 37℃for 72 hours. On day eight, the cell culture plates were removed and centrifuged at 300g for 3 minutes to collect 200. Mu.l/well of culture supernatant. HBV DNA extraction in cell culture supernatants was performed using Qiagen automated DNA extraction equipment, specific methods being referred to reagents and instrument instructions. Finally, the extracted DNA was eluted with DNA elution buffer at 100. Mu.l/well. The HBV DNA quantitative PCR analysis is carried out on the extracted DNA by adopting a quantitative detection kit for the hepatitis B virus nucleic acid of the Taipu organism, and the specific method is described by referring to the kit. The quantitative standard curve adopts a kit to carry a standard sample, and experiments are carried out in parallel. Each sample was quantitatively converted according to a standard curve. Finally, using Graphpad Prism software to calculate the EC of the compound according to each concentration of the compound and the corresponding DNA value ₅₀ Values. Emax is the effect value of the compound to maximally inhibit HBV DNA replication.

Compounds of the present disclosure inhibit HBV capsid protein assembly byFurther, the in vitro activity of inhibiting HBV DNA replication was determined by the above test, and the EC was determined ₅₀ The values are shown in Table 1.

TABLE 1 EC of anti-HBV activity test (intracellular HBV DNA quantitative analysis) of the compounds of the present disclosure ₅₀ Value of

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Conclusion: the compound has obvious inhibition effect on HBV DNA replication.

Test example 2, effect of the compounds of the present disclosure on proliferation of HepG2 cells in vitro

1. Experimental material and instrument

HepG2 cells (ATCC)

2.CellTiter-Glo ^TM Cell proliferation kit (Promega)

3. Automatic pipetting station (Bravo): agilent Technologies Co Ltd

4. Microplate reader (VICTOR 3): perkinelmer Co

5.CO ₂ Incubator (Fisher Scientific)

6. Centrifuge (Fisher Scientific)

2. Experimental procedure

HepG2 cells in the logarithmic growth phase were taken, and cell suspensions were prepared by trypsin digestion, and incubated in 6,000 cells/well 96-well plates (bottom-clear white 96-well plates, perkinelmer) with 5% carbon dioxide at 37℃for 16-20 hours. The next day, compounds were dissolved in pure DMSO at a concentration of 20mM, and were serially diluted 3-fold using an automated pipetting station (Bravo), with 8 concentration points per compound, and control wells in DMSO; each concentration point compound in DMSO was then 200-fold diluted with EMEM (10% fbs in) medium. Taking out the cell culture plate inoculated on the first day, sucking out the culture medium in the pore plate by using a negative pressure sucking device, and then mixing the prepared culture medium with various concentrations The compound culture medium was added to each well and incubated at 37℃for 72 hours at 100. Mu.l/well. On the fifth day, 96-well cell culture plates were removed, freshly prepared CellTiter Glo was added to each well, left for 5-10 minutes at 100 μl/well, the bottom of the 96-well plate was sealed with a white back cover (PerkinElmer), placed in an microplate reader, and the Luminescence signal was measured with the microplate reader. Calculation of CC for the Compound from the respective concentrations of the Compound and the corresponding proliferation inhibition Signal values Using Graphpad Prism software ₅₀ Values.

The effect of the compounds of the present disclosure on inhibition of proliferation of HepG2 cells in vitro was determined by the above assay, CC measured ₅₀ The values are shown in Table 2.

TABLE 2 CC of the compounds of the present disclosure for inhibition of proliferation of HepG2 cells in vitro ₅₀ Value of

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Conclusion: the compound disclosed by the disclosure has no or little influence on the inhibition of proliferation of HepG2 cells in vitro and shows high safety.

Pharmacokinetic evaluation

Test example 3 pharmacokinetic testing of the presently disclosed compounds

1. Summary

The drug concentration in plasma was measured in mice as test animals by LC/MS method at various times after the compounds of examples 2, 6, 13 were administered by intragastric administration. Pharmacokinetic behavior of the compounds of the present disclosure in mice was studied and their pharmacokinetic profile was assessed.

2. Test protocol

2.1 test drug

Example 2, example 6, example 13.

2.2 test animals

The C57 mice, 27 females, were equally divided into 3 groups of 9 animals each, purchased from velariwa laboratory animal technologies ltd, animal production license number: SCXK (thunberg) 2019-0001; animal pass number: 1907080105.

2.3 pharmaceutical formulation

A certain amount of medicine is weighed, and 5% of DMSO, 5% of Tween 80 and 90% of physiological saline are added to prepare a clear and transparent solution of 0.1 mg/mL.

2.4 administration of drugs

One group of mice is administrated by stomach irrigation, the administration dosage is 2.0mg/kg, and the administration volume is 20.0mL/kg.

3. Operation of

Mice were perfused with the compounds of examples 2, 6, 13, and 0.1mL of blood was collected from the orbit before and after administration for 0.25,0.5,1.0,2.0,4.0,6.0,8.0, 11.0, 24.0 hours, placed in heparinized tubes, and centrifuged at 10000rpm for 5 minutes at 4℃to separate plasma, which was stored at-20℃and fed 2 hours after administration.

Determining the content of a compound to be detected in the plasma of a mouse after the gastric lavage administration of the drug: 25. Mu.L of mouse plasma at each time after administration was taken, 50. Mu.L (100 ng/mL) of camptothecin as an internal standard solution, 200. Mu.L of acetonitrile, vortex-mixed for 5 minutes, centrifuged for 10 minutes (4000 rpm), and 1.0. Mu.L of the supernatant was taken from the plasma sample for LC/MS/MS analysis.

4. TABLE 3 pharmacokinetic parameter results

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Conclusion: the compound disclosed by the disclosure has the advantages of good drug absorption, good bioavailability and pharmacokinetic advantage.

Test example 4 liver exposure/liver plasma ratio of the compounds of the present disclosure

1. Summary

The liver exposure/liver plasma ratio was determined 1 hour after gavage administration in rats as the test animals.

2. Test protocol

2.1 test drug

Example 6 and example 13.

2.2 test animals

Healthy adult SD rats, 8, were purchased from vitelliwa laboratory animal technologies, inc, in two groups, male and female halves, animal production license number: SCXK (Zhe) 2019-0001, certification No. 1909050050.

2.3 pharmaceutical formulation

A certain amount of medicine is weighed, and 0.1 percent of Tween 80 and 99.9 percent of 1 percent of HPMC are added to prepare a uniform suspension solution with the concentration of 2 mg/mL.

2.4 administration of drugs

SD rats were dosed by gastric lavage overnight fast with a dose of 20mg/kg of the compound of example 6, 20mg/kg of the compound of example 13, and a dosing volume of 10mL/kg.

3. Operation of

The compound of example 6 and the compound of example 13 were administered by gavage to rats, and the orbital blood was collected (anticoagulation with heparin sodium, 0.5 ml) 1 hour after administration, and centrifuged at 10000rpm for 1min (4 ℃) and stored at-20 ℃. The abdominal aorta was exsanguinated after 20% uratam anesthesia in rats, the blood was collected, the liver was isolated and washed several times with physiological saline, and the water was sucked by filter paper and stored frozen at-20 ℃.

Determination of the content of the test compound in the plasma and liver of rats 1 hour after administration of the drug by gastric lavage: rat plasma or liver homogenate (10 times volume PBS homogenate) 25. Mu.L after 1 hour of administration was taken, 50. Mu.L (100 ng/mL) of the internal standard solution camptothecin, 200. Mu.L of acetonitrile, vortexing for 5 minutes, centrifuging for 10 minutes (4000 rpm), and 2.0. Mu.L of the supernatant was taken and analyzed by LC/MS/MS.

4. Pharmacokinetic parameter results

Table 4 liver exposure/liver plasma ratios of compounds of the present disclosure are as follows:

conclusion: the liver exposure/liver plasma ratio is high, and the tissue distribution is good.

Test example 5 inhibition of human liver microsome P450 subunit enzyme CYP2C9,1A2,2D6,3A4T,2C19 by Compounds of the disclosure

The enzymatic activity of the compounds of the present disclosure on human liver microsome P450 subunit CYP2C9,1a2,2d6,3a4t,3a4m,2C19 was determined using the following experimental method.

1. Experimental material and instrument

1. Phosphate buffer (20 XPBS, purchased from Producer)

2.NADPH(ACROS，A0354537)

3. Human liver microsomes (Corning Gentest, cat No.452161, lot No.6123001, 33 Donors)

ABI QTrap 4000 liquid and mass dual-purpose instrument (AB Sciex)

Inertsil C8-3 column, 4.6X105 mm,5 μm (America Dimma Co.)

CYP Probe substrate 2C9 (diclofenac/4. Mu.M, powder from SIGMA, cat No. D6899-10G), 2C19 ((S) -mephenytoin/20. Mu.M, powder from carboline technologies Co., ltd., cat No. 303768); 1A2 (phenacetin/12. Mu.M, powder from China medicine biosciences institute, 100095-200204), 2D6 (dextromethorphan/4. Mu.M, powder from SIGMA, cat No. D9684-5G) and 3A4T (testosterone/75. Mu.M, powder from Wobbe, cat No. 58-22-0); positive control inhibitors 2C9 (sulfabenazole, powder from SIGMA, cat No. 526-08-9), 1A2 (α -naphthaleneflavone, powder from SIGMA, catNo.N5757-1G), 2D6 (quinidine, powder from SIGMA, cat No. q0750-5G), 3A4T (ketoconazole, powder from SIGMA, cat No. k1003-100 MG) and 2C19 (ticlopidine, powder from SIGMA, cat No. T6654-1G).

2. Experimental procedure

100mM PBS buffer was used to prepare 2.5mg/mL microsomal solution and 15mM MgCl ₂ And 5mM NADPH solution, with DMSO the stock solution having a concentration of 30mM was diluted to a series of solutions I having a concentration of 10mM, 3mM, 1mM, 0.3mM, 0.03mM, 0.003mM, and 0mM, and the series of solutions I was further diluted 200 times with Phosphate Buffer (PBS) to obtain a series of solutions II (150, 50, 15, 5, 1.5, 0.15, 0.015, 0. Mu.M) to be tested.

2.5mg/mL of microsomal solution, 20. Mu.M diclofenac (2C 9) or 60. Mu.M phenacetin or 20. Mu.M dextromethorphan or 375. Mu.M testosterone or 100. Mu.M ((S) -merphenytoin (2C 19) working solution, mgCl were taken, respectively ₂ The solutions and compound working solutions (150, 50, 15, 5, 1.5, 0.15, 0.015, 0. Mu.M, corresponding reaction systems were set for each concentration) were each 20. Mu.L, and mixed well. The positive control group replaced the compound with the same concentration of sulfabenzpyrazole (2C 9), α -naphthaleneflavone (1 A2), quinidine (2D 6), ketoconazole (3 A4T) or ticlopidine (2C 19). While 5mM NADPH solution was preincubated together at 37℃for 5 minutes. After 5 minutes 20. Mu.L of NADPH was added to each well, the reaction was started and incubated for 30 minutes. All incubation samples were double-sampled. After 30 minutes 200. Mu.L of acetonitrile containing internal standard was added to all samples, mixed well, shaken at 800rpm for 10 minutes, and centrifuged at 3700rpm for 10 minutes. 100. Mu.L of the supernatant was mixed with 80. Mu.L of water and analyzed by LC-MS/MS.

The numerical value is calculated by Graphpad Prism to obtain the IC of the medicine to CYP2C9 diclofenac, 1A2 phenacetin, 2D6 dextromethorphan, 3A4T testosterone and 2C19 (S) -mefenadine metabolic sites ₅₀ The values are shown in Table 5.

TABLE 5 IC of the presently disclosed compounds for CYP2C9 diclofenac, 1A2 phenacetin, 2D6 dextromethorphan, 3A4T testosterone, and 2C19 (S) -mefenadine metabolic sites ₅₀

Conclusion: the disclosed compounds have no inhibition on human liver microsomal CYP2C9 diclofenac and 2C19 (S) -mefenadine metabolic sites, exhibit better safety in terms of drug interactions, suggesting that no metabolic drug interactions based on inhibition of CYP2C9 diclofenac and 2C19 (S) -mefenadine metabolic sites by the compounds occur.

Test example 6 blocking of hERG potassium current by the compounds of the present disclosure

1) Purpose of experiment

The blocking effect of the compounds of the present disclosure on hERG potassium current was tested on stable cell lines transfected with hERG potassium channels using fully automatic patch clamp.

2) Experimental method

2.1 Experimental materials and instruments

Experimental materials:

experimental instrument:

2.2 Full-automatic patch clamp experimental step

HEK293-hERG stable cell lines (according to known technology built-in) in DMEM/HIGH glucose medium (10% FBS,1.5 u g/ml puromycin dihydrochloride) at a density of 1:4 for 48-72 hours in full automatic patch clamp experiments. After cells were digested with 0.25% pancreatin on the day of the experiment, cells were collected by centrifugation and resuspended in extracellular fluid to make a cell suspension. The cell suspension was placed on the cell bank of a Patchliner instrument which applied cells to the chip (NPC-16) using a negative pressure controller, and the instrument attracted individual cells to the wells of the chip by negative pressure. After the whole cell mode is established, the instrument will obtain hERG current according to the set hERG current voltage program, and then the instrument automatically performs compound perfusion from low concentration to high concentration. The current at each concentration of compound was analyzed as well as the blank current by data analysis software provided by HEAK Patchmaster, HEAK EPC10 patch clamp amplifier (Nanion) and Pathlinessoftware and Pathcontrol HT software.

2.3 Test results

Compounds of the present disclosure are directed against hERG potassium currentThe blocking effect of (2) was measured by the above test, and the measured IC ₅₀ The values are given in Table 6 below.

TABLE 6 IC of blocking of hERG potassium currents by compounds of the present disclosure ₅₀

Examples numbering	IC ₅₀ (μM)
		1	1.3
6	11
		13	18

Conclusion: the compounds of the present disclosure have significantly improved inhibition of hERG when they introduce carboxylic acid groups into the molecule.

Claims

1. A compound of the general formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof,

wherein:

ring a is a 6 to 10 membered aryl;

L ¹ is C _1-6 An alkylene group;

R ¹ is C _1-6 An alkyl group;

R ² identical or different and are each independently selected from hydrogen atoms, halogenElement, C _1-6 Alkyl and C _1-6 A haloalkyl group;

R ³ and R is ⁴ Forming a 3-to 6-membered heterocyclic group with the nitrogen atom to which it is attached, said 3-to 6-membered heterocyclic group optionally containing 1 to 2 hetero atoms which are the same or different and are selected from N, O and S, in addition to 1 nitrogen atom, and said 3-to 6-membered heterocyclic group optionally being further selected from C _1-6 Alkyl, halogen, C _1-6 Haloalkyl, C _1-6 Alkoxy, C _1-6 Haloalkoxy, hydroxy, C _1-6 Hydroxyalkyl, 3-to 6-membered cycloalkyl, 3-to 6-membered heterocyclyl and-L ² -C(O)OR ⁹ Is substituted by one or more substituents;

R ⁵ identical or different and are each independently selected from hydrogen atoms, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy and C _1-6 Haloalkoxy groups;

R ⁶ identical or different and are each independently selected from hydrogen atoms, C _1-6 Alkyl, halogen and C _1-6 A haloalkyl group;

R ⁹ selected from hydrogen atoms, C _1-6 Alkyl and C _1-6 A haloalkyl group;

L ² selected from the group consisting of a bond, a 3-to 6-membered cycloalkyl group, and C _1-6 Alkylene, said 3-to 6-membered cycloalkyl and C _1-6 Alkylene is optionally further selected from C _1-6 Alkyl, halogen, C _1-6 Haloalkyl, C _1-6 Alkoxy and C _1-6 One or more substituents in the haloalkoxy group;

r is selected from 0, 1 or 2;

s is selected from 0, 1, 2, 3 or 4; and is also provided with

t is selected from 0, 1, 2, 3, 4 or 5.

2. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein L ¹ Is CH ₂ 。

3. A compound of formula (I) according to claim 1, wherein ring a is phenyl, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof.

4. The compound of formula (I) according to claim 1, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, which is a compound of formula (II), or a diastereomer thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ¹ to R ⁶ R, s and t are as defined in claim 1.

5. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein R ⁶ Is a hydrogen atom.

6. The compound of formula (I) according to claim 1, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, which is a compound of formula (III):

wherein:

R ¹ to R ⁵ S and t are as defined in claim 1.

7. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein R ³ And R is ⁴ Forming a 5-to 6-membered heterocyclic group with the linking nitrogen atom; the 5-to 6-membered heterocyclic group optionally contains 1 to 2 identical or different heteroatoms selected from N and O in addition to 1 nitrogen atom, and the 5-to 6-membered heterocyclic group is optionally further substituted by one or more-L ² -C(O)OR ⁹ Substituted;

L ² selected from the group consisting of a bond, a 3-to 6-membered cycloalkyl group, and C _1-6 Alkylene, said 3-to 6-membered cycloalkyl and C _1-6 Alkylene groups optionally further being substituted by one or more C' s _1-6 Alkyl substituted;

R ⁹ is a hydrogen atom or C _1-6 An alkyl group.

8. A compound of formula (I) according to claim 1, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, which is a compound of formula (IV):

wherein:

R ⁷ identical or different and are each independently selected from C _1-6 Alkyl, halogen, C _1-6 Haloalkyl, C _1-6 Alkoxy, C _1-6 Haloalkoxy, hydroxy and C _1-6 A hydroxyalkyl group;

R ⁸ is a hydrogen atom OR-C (O) OR ⁹ ；

R ⁹ Is a hydrogen atom or C _1-6 An alkyl group;

u is selected from 0, 1, 2, 3, 4 or 5;

R ¹ 、R ² 、R ⁵ s and t are as defined in claim 1.

9. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein R ⁵ Is C _1-6 Alkyl or halogen。

10. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, which is a compound of formula (V) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof:

wherein:

R ^5a and R is ^5b Identical or different and are each independently selected from C _1-6 Alkyl, halogen, C _1-6 Haloalkyl, C _1-6 Alkoxy and C _1-6 Haloalkoxy groups;

u is selected from 0, 1, 2, 3 or 4;

R ¹ 、R ² 、R ⁷ 、R ⁸ 、L ² and s is as defined in claim 8.

11. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 10, wherein R ^5a And R is ^5b Identical or different and are each independently selected from C _1-6 Alkyl or halogen.

12. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 10, wherein R ⁷ Is a hydrogen atom or C _1-6 An alkyl group.

13. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 10, wherein R ⁷ Is a hydrogen atom.

14. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 8, wherein R ⁸ is-C (O) OH.

15. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein L ² Is a key.

16. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein L ² Is 3-6 membered cycloalkyl or C _1-6 Alkylene, said 3-to 6-membered cycloalkyl or C _1-6 Alkylene groups optionally further being substituted by one or more C' s _1-6 Alkyl groups are substituted.

17. The compound of formula (I) or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, according to claim 1, wherein R ² Is a hydrogen atom.

18. A compound of formula (I) according to claim 1, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, selected from:

19. A compound as shown below, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, selected from:

20. a process for preparing a compound of formula (I) according to claim 1 or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, which comprises the steps of:

wherein X is halogen; ring A, R ¹ To R ⁶ 、L ¹ R, s and t are as defined in claim 1.

21. A pharmaceutical composition comprising a therapeutically effective amount of a compound of general formula (I) according to any one of claims 1 to 18, or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.

22. Use of a compound of general formula (I) according to any one of claims 1 to 18 or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 21, for the preparation of a capsid protein inhibitor.

23. Use of a compound of general formula (I) according to any one of claims 1 to 18 or a diastereomer thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 21, for the manufacture of a medicament for the treatment of a viral infection disorder.

24. The use according to claim 23, wherein the virus is selected from the group consisting of hepatitis b virus, influenza virus, herpes virus and aids virus and the disease is selected from the group consisting of hepatitis b, influenza, herpes and aids.